Progress 10/01/08 to 09/30/13

Outputs
Target Audience: This past year we gave 3 presentations (1 oral and 2 posters) at the Institute of Food Technologists Annual Meeting in Chicago . The oral was attended by over 100 scientists; graduate students and faculty from around the world, and industry scientists who worked on protein functionality (both physical/chemical and bioactivity). We delivered a copy of the posters to at least 50 of these scientists. In addition we collected their business cards and sent them an email with a link to Dropbox where we have in one folder all the presentations given and papers published since we started this work on protein aggregation in 2008. In addition we presented 2 posters at the NIFA PI meeting attended by about 39 faculty who were awarded NIFA grants. We also gave 2 oral presentation to Davisco International Inc, who are one of the major producers of whey isolates in the world, and who gave us supplemental funding in the area of the work on glycation. In addition I attended both the Chicago and Minnesota IFT sections Suppliers Day meeting where there were at least 200 and 150 table top exhibitors, respectively. I delivered info on our functionality work to all the protein exhibitors I could find (about 45 in total). We have published refereed research papers in the Journal of Food Science, Journal of Agricultural & Food Chemistry, Journal of Raman Spectroscopy, Food Research International and Food Chemistry (From 2008 until present we have published 6 papers on dry and intermediate moisture (IMF) model protein bar systems focusing on egg white and eg yolk and 11 papers focusing on storage stability of whey proteins, casein and their hydrolysates under temperature abuse at different moisture contents. . In addition we have a review paper published in Critical Reviews in Food Science and Nutrition which has an impact factor of 4.8. The paper is a review focusing on storage stability of food protyein hydrolysates, and was commissioned by the Midwest Dairy Center. It should be noted that we are collaborating on this work with Dr. Peng Zhou of Jingnan Univ. in Wuxi PRC. He has given at least 5 lectures in China on our combined work. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? This project has supported the work of two visiting scientists: John Carlo Rocca (Univ. Udine in Italy) and Andre Klasses (Univ. Waganingen, Netherlands). We trained 6 MS students, over the grant period, 3 will be finishing their thesis workin the Spring of 2014. The PhD student will be done by June 2014. Our collaborator, Dr. Zhou had 5 MS and 1 PhD student involved in the work at Jiangnan Univ. The project also partially supported 6 postdocs. How have the results been disseminated to communities of interest? As noted earlier we had made presentations as either posters or oral presentations every year at the national annual meeting of the Institute of Food Technologists. In every case we had prepared copies of the posters and Power Point to those of interest which included faculty, grad students and industry researchers. I estimate that our findings have been presented to at least 500 people over the past years. The earlier research papers have had much exposure so already have many citations. Some examplesare listed here. It should be noted that the Times Higher Education website latest information indicated that in 2007 in the agricultural sciences, which would include food science, the average citation rate per paper averaged over 10 years (2000 to 2010) was 7 with those published since 2007 were 5.24, thus our papers are being considered important. Effect of water content on glass transition and protein aggregation ofwheyprotein powders during short-term storage P Zhou,TPLabuza- Food Biophysics, 2007 - Springer Abstract The objectives of this study were to investigate the moisture-induced protein aggregationofwheyprotein powders and to elucidate the relationship of protein stability with respect to watercontent and glass transition. Threewheyprotein powder types were studied:wheyprotein... Cited by 39Related articlesAll 3 versionsWeb of Science: 26CiteSave Moisture-induced aggregation ofwheyproteins in a protein/buffer model P Zhou, X Liu,TPLabuza- Journal of agricultural and food …, 2008 - ACS Publications Moisture-induced protein aggregation in a dry or intermediate-moisture food matrix can contribute to the loss of product acceptability. The present study evaluated the molecular mechanisms and controlling factors for moisture-inducedwheyprotein aggregation in a... Cited by 29Related articlesAll 4 versionsWeb of Science: 21 Effects of moisture-inducedwheyprotein aggregation on protein conformation, the state of water molecules, and the microstructure and texture of high-protein- … P Zhou, X Liu,TPLabuza- Journal of agricultural and food …, 2008 - ACS Publications Moisture-induced protein aggregation through intermolecular interactions such as disulfide bonding can occur in a high-protein-containing food matrix during nonthermal processing and storage. The present study investigated the effect of moisture-inducedwheyprotein... Cited by 29Related articlesAll 3 versionsWeb of Science: 23CiteSave Effects of polyols on the stability ofwheyproteins in intermediate-moisture food model systems X Liu, P Zhou, A Tran,TPLabuza- Journal of agricultural and food …, 2009 - ACS Publications The objective of this study was to investigate the influence of polyols on the stability ofwhey proteins in an intermediate-moisture food model system and to elucidate the effect of polyols on the hardening ofwheyprotein-based bars during storage. Four major polyols, glycerol,... Cited by 15Related articlesAll 4 versionsWeb of Science: 10 In addition to the above, presentations on protein aggregation were also made at the Euro Foods Water meetings in Helsinki Finland and Stutgard Germany (over 120 at each meeting) and to the International Whey Conferences held in Chicago Illinois and in Paris France with over 200 at each meeting. In addition we have made several presentations at Davisco International in Minnesote who are the largest producers of isolated whey in the USand have discussed our findings with researchers from Glanbia (Idaho) and 3 protein specialty companies in Minnesota (Vision Processing (Owatonna); Sun Opta (Cambridge) ;and Milk Specialties (Eden Prairie). What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? With respect to Goal 1, we decided to focus only on bioactive proteins and their hydrolysates, the latter which have biological health functionality, e.g. reduction of high blood pressure. Preliminary work in 2006-08 indicated that significant protein-protein aggregation occurs during storage causinginsolubility as well as the hardening of the high protein nutritional bar. This would lead to loss of palatability. Thus our focus turned to looking at the cause of this and ways to prevent occurrence. This was also driven by the receipt of a $500,000 NIFA Grant,a $300,000 grant from the American Egg Board, $50,000 support from Davisco and support from the Midwest Dairy Center. For Goal 2 we were able to partner with Dr. Tonya Schoenfuss, in our department who has a capillary electrophoresis HPL for separations by molecular weight. We also used SDS PAGE to determine dimer formation of the proteins. For Goal 3 we made protein bars using a commercial; formula using different humectants, different proteins and their hydrolysates and glycated proteins , the later being carried out in part by Dr. Zhou in China where he confirmed structural changes by mass spectrometry and by Dr. Pam Ismail of our department who confirmed aggregation by Raman Spectroscopy. These were stored at room temperature, 35°Cand 45°C. Aggregation was assessed by texture analysis and confirmed by SDS PAGE. In addition we measured color (darkness and available amines as indices for the Maillard reaction which can also lead to aggregation. For Goal 4 we found several factors that caused hardness and ways to reduce or eliminate it. One simple way is to reduce the moisture to below the BET monolayer value. . This stops all water phase reactions including aggregation but limits the protein delivery to zero. Since most protein peptides are bitter, they are unpalatable. A second approach is to glycate the proteins with a short chain oligomer (10 to 20) dextran. The dextran is chopped off in the small intestine thereby getting back its health benefit. Thus we had to key the format as an IMF soft food bar. In these systems aggregation still occurred through disulfide bond formation when no reducing reducing sugars were used. Unfortunately most disulfide bond compounds are highly toxic. Cystiene is somewhat beneficial but imparts a sulfury taste/aroma which is objectionable. Using reducing sugars like high fructose corn syrup (HFCS), the bar acceptability was even worse as it also induced the Maillard browning reaction which also causes protein aggregation through other covalent bonding. Using maltitol in place of HFCS significantly reduced the Maillard reaction but not to zero because of residual glucose in the commercial product. We also found that egg white protein also browned even though it is treated with glucose oxidase toreduce glucose levels below 1%. We also found that propylene glycol caused rapid aggregation even at low moistures, the mechanism needs further research. Thus a good formula uses glycerol and maltitol (similar sweetening power as sucrose) as the humectants to control water activity but disulfide bonding still occurs. To finally solve this we used a combination of both intact and hydrolyzed proteins. The latter have glass transitions at much lower temperature that the corresponding intact protein. With about 20 % substitution with the hydrolysates the bars remained soft and had an acceptable texture for at least 9 months at room temperature so we met our goal. Also applying the kinetics to the hardening data showed that Accelerated Shelf Life Testing (ASLT) at 45°C increased the reaction rate significantly with a factor of 10 to 11x faster than at 23C so that industry can use this to shorten product development time. Because of lack of funding with respect to Goals 5 and 6 and the cost of running the extruder in our pilot plant at about $1500 a day, we did not pursue this route. Goal 7 is covered by this final report.

Publications

• Type: Journal Articles Status: Accepted Year Published: 2013 Citation: Zhou, P., Guo, Mufan, Liu, D., Liu, X., and Labuza, T.P. 2013. Maillard Reaction-Induced Modification and Aggregation of Proteins and Hardening of Texture in a Protein Bar Model System. J. Food Science 78(3) C437-C444; 2013 http://dx.doi.org/10.1111/1750-3841.12061 Qian Wang, Lili He, Theodore P. Labuza and Baraem Ismail. 2013. Structural Characterization of Partially Glycosylated Whey Protein as influenced by pH and Heat using Surface-Enhanced Raman Spectroscopy. Food Chem. 139 (1-3):313-319 http://dx.doi.org/10.1016/j.foodchem.2012.12.050 Rao, Q., Klassen, A., and Labuza, T.P. 2013 Storage stability of Food Protein Hydrolysates. Critical Reviews in Food Science and Nutrition. DOI http://dx.doi.org/10.1080/10408398.2012.758085 Lili He, Jinkai Zheng, Theodore P. Labuza and Hang Xiao. 2013 A surface enhanced Raman spectroscopic study of interactions between casein and polymethoxyflavonesJ Raman Spectroscopy 44 (4) 531-535 DOI: http://dx.doi/10.1002/jrs.4229 Rao, Qinchun, Fisher, Mary, Gao, Mufan, and Labuza, Theodore P.; 2013. Storage stability of a commercial hen egg yolk powder in dry and intermediate-moisture food matrices. J. Agri. Food Chemistry 61 (36), pp 8676-8686; DOI: http://pubs.acs.org/doi/abs/10.1021/jf402631y? Yingjiq, Chen, Xiaoming;Liu, Theodore Labuza; Peng Zhou. Effect of molecular size and charge state of reducing sugars on nonenzymatic glycation of B-lactoglobulin. Food Research International. 2103. 54: 1560-1568. DOI:http://dx.doi.org/10.1016/j.foodres.2013.09.030 Peng Zhou, Dasong Liu, Xiaoxia Chen, Yingjia Chena and Theodore P. Labuza Stability of whey protein hydrolysate powders: effects of relative humidity and temperature 2013 Food Chemistry http://dx.doi.org/10.1016/j.foodchem.2013.027 Accepted

Progress 01/01/12 to 12/31/12

Outputs
OUTPUTS: The main focus this year was in 3 areas. The first was the continuation of the study of protein aggregation of various commercial dry protein ingredients which are used in nutritional high protein bars for weight management/muscle recovery/meal substitution. Prior work showed that high protein whey bars become harder over time without moisture loss. This allowed us to obtain funding from the American Egg Board (AEB), Davisco Foods and a USDA NIFA grant for 2012-13. We focused on dry egg white, hydrolyzed white and dry egg yolk. Measurement of aggregation was by SDS Page, use of disulfide bond breakers, use of capillary electrophoresis liquid chromatography for size separation as well as texture hardness, color change, fluorescence and the OPA method for available amines (Maillard reaction - NEB). Also moisture sorption (m) isotherms, monolayer and Tg as f(m) were determined .Model systems of different composition and water activities were studied at 3 temperatures to establish Q10 values (rate increase for 10C temperature increase). Results were presented at: IFT annual Mtg. (3 posters, 1 oral); USDA-NIFA PI Mtg., Min. Dairy Assoc. Annual Mtg., Davisco, Euro Foods Water Mtg. in Helsinki, Dept. FScN 40th celebration with 250+ attendees (2 posters) , and copies of both the posters and published papers (5 this year) were distributed. An Executive Summary on dry egg powders including a packaging prediction spreadsheet was distributed to the AEB which comprises many egg processors. Area 2 was continuation of evaluation of moisture re-distribution in trail mix products funded by the Almond Board of CA (ABC). Almonds were stored in jars with other ingredients (eg dried fruit, choc-chips etc.) and moisture changes with time measured and compared to predictions so as to determine a "best if used by date". A final report was distributed to the ABC board who distributed it to the processors. The 3rd area was to establish a new platform for rapid analysis of adulterants (<30 min) in food including bioterror agents (ricin, anthrax; Dept. Homeland Security grant), allergens (milk/egg, Dairy Research Inc. grant) and pesticides on produce (USDA-NIFA grant co-PI grant with Vet Med., Chemistry and Food Sci.) . The protocol used is to capture the adulterant by an antigen/antibody reaction with the or by a single stranded DNA aptamer. Aptamers were created with Dept. Vet Med collaboration using the Select system of DNA databases. Either the aptamer or antibody is bonded onto a magnetic bead and is used to capture and separate the target from a food suspension followed by releasing the adulterant with deposition on a silver dendrite nano-surface, then measured by Raman spectroscopy using PCA.. Presentations were made at IFT, and IFPA meetings and at several companies including Neogen. In addition we have presented 3 Thermo-Fisher webinars and 3 NCFPD webinars on this platform. Thermo Fisher has donated a state of the art RS instrument. Two undergrads scholarships based on this work from DHS The Royal Chem. Soc. has deemed this cutting edge work and accepted and published papers in Analyst and Chemical Science in 35 days of receipt. PARTICIPANTS: Individuals Prof. Theodore P. Labuza (Co-PI) Dept. Food Science & Nutrition, Univ. Minnesota Assoc. Prof. Baram Ismail (Co-PI) Dept. Food Science & Nutrition, Univ. Minnesota Asst. Prof Lili He (Co-PI) Dept. Food Science & Nutrition, Univ. Minnesota and Univ. of Massachusetts Asst. Prof. Stephan Lumor (Co-PI) Dept. Food Science & Nutrition, Univ. Minnesota & Delaware State Univ. Prof. Francisco Diez-Gonzalez Dept. Food Science & Nutrition, Univ. Minnesota Prof Srinand Sreevatsan (Co-PI) Dept Veterinary Medicine Univ. of Minnesota Assoc. Prof. Chisty Haynes (Co-PI) Dept of Chemistry Univ. of Minnesota Prof Peng Zhou (collaborator) Dept. of Food Science Jingnan Univ. Wuxi, China Dr. Qinchun Rao Senior Research Associate, Dept Food Science and Nutrition, Univ. of Minnesota, Tom Rodda MS Student Dept. Food Science & Nutrition, Univ. Minnesota Mufan Guo PhD student Dept. Food Science & Nutrition, Univ. Minnesota Tuo Chen MS student, Dept. Food Science & Nutrition, Univ. Minnesota Jean Carlo Rocca MS student Univ. Udine Italy and Dept. Food Science & Nutrition, Univ. Minnesota Andre Klassen Visiting Scientist Univ. Wageningen Netherlands & Dept. Food Science & Nutrition, Univ. Minnesota Allysa Pagel Undergrad Dept. Food Science & Nutrition, Univ. Minnesota Ian Ronnigan Undergrad, Dept. Food Science & Nutrition, Univ. Minnesota Neal Fredrickson Undergrad, Dept. Food Science & Nutrition, Univ. Minnesota Ken Smith Undergrad Dept.of Chemistry St. Scholastica Duluth Minnesota Partner Organizations Thermo Fisher Scientific TARGET AUDIENCES: Food processing companies Government Food Safety Organization including FSIS-USDA, CFSAN-FDA, EPA, DHS, State Health Departments, other researchers in universities and USDA Labs PROJECT MODIFICATIONS: Not relevant to this project.

Impacts
Area 1: We were able to discern 3 reaction modes causing protein aggregation/hardening and loss of solubility/nutritional value during storage of intermediate moisture protein bars. All modes increased in rate as storage water activity and temperature increased. These modes include: (1) Hydrophobic/Hydrophilic bonding between proteins. These are weak bonds with low energy so low buffer concentrations will break the bonds allowing solubilization and maintenance of protein nutritional quality during digestion; (2) Inter-protein covalent disulfide bond formation. These are irreversible bonds under normal conditions. Adding cysteine at low concentrations (~0.1%) slows bond formation; (3) Intra and inter molecular covalent bonding as the result of the Maillard reaction between reducing sugars and amine groups. This mode of aggregation is also irreversible. One way to prevent the Maillard reaction is to remove any residual glucose from dry egg powders and lactose from whey powders. A second is to use sugar alcohols in place of corn syrup or high fructose corn syrup in protein bar formulations. A third is to react the proteins with dextrans at controlled conditions which protects the protein from further reaction and yet is digestible. These studies also suggest that, at least in dry egg yolk powder (DEY), oxidation of unsaturated lipids causes free radical damage to the protein resulting in aggregation and thus loss of solubility. Some approaches for maintaining a soft desirable texture for protein bars include substituting up to 25% of the protein in the formula with a 5 to 7% hydrolyzed protein (e.g. hydrolyzed whey or egg white). This is the result of a significant reduction in glass transition temperature. Alternatively, one may substitute ~25% of the protein with DEY since DEY maintains softness because of its 50% fat content. A final alternative is to determine the optimum moisture contents so as to maximize the quality of the powders during distribution before being incorporated in final products. There were 9 refereed journal articles published or in press. Three of them were resulted from collaborative work at Jingnan Univ. in Wuxi, China. In area 2 it was found that the moisture exchange between almonds and other ingredients took place in about 2-3 weeks and the equilibrium water activity could be predicted from a composite isotherm. In area 3, the development of IMS-SERS, we continued to optimize and refine the methods. Three papers were published in 2012. We were able to successfully detect anthrax spores at 10^3/mL from water, milk and orange juice using published aptamer structures. A toxic dose is ~10^6 spores so this is adequate for rapid testing. We found that we could not create useful aptamers for water insoluble pesticides but direct methanol swabs of a steel surface and from an apple surface showed success with a LOD for Thiabendazole of 10 ppb. The method easily could differentiate a mixture of DDT, Dieldrin and malathion using PCA. For allergens, using an aptamer for egg ovalbumin, we were able to detect it in milk @ 2 micro-g/mL

Publications

• Referred Journal Articles (2012)
• Rao, Q, Rocca-Smith, J.R., and Labuza, T.P. (2012) Moisture induced Quality Changes of Hen Egg White proteins in a Protein/water model system. J. Agr. Food Chem. 60(42): 10625-10633. DOI:10.102/jf302402k l
• Rao, Q., and Labuza, T. P. (2012) . Effect of moisture content on selected physicochemical properties of two commercial hen egg white powders. Food Chemistry 132:373-384 doi:10.1016/j.foodchem.2011.10.10
• Rao, Q, Rocca, J.C., Shoenfuss,T. and Labuza, T.P., (2012) Accelerated shelf-life testing of quality loss for a commercial hydrolyzed hen egg white powder. 2012 Food Chemistry 135 (2): 464-472
• Liu, D., Zhou, P., Liu, X., and Labuza, T.P., (2012) Moisture-induced aggregation of alpha Lactalbumin: Effects of temperature, cations and pH. J. Food Science 76(6): C817-C823. DOI: 10.1111/j.1750-3841.2011.02277.x
• Chen, Y, Liang, L., Liu, X., Labuza, T.P., and Zhou, P. (2012) Effect of Fructose and Glucose on Glycation of Beta-Lactoglobulin in an Intermediate Moisture Food Model System: Analysis by Liquid Chromatography Mass Spectrometry (LC MS) and Data Independent Acquisition LCMS, (LC MS^E). J. Agric. Food Chem., 60 (42), pp 10674-10682 DOI: 10.1021/jf3027765
• Rao, Q., Rocca-Smith, J., Labuza. (2012) Storage stability of hen egg white powders in three protein/water dough model systems. Food Chemistry (accepted in press)
• Zhou, P., Guo, Mufan, Liu, D., Liu, X., and Labuza, T.P. (2012) Maillard Reaction-Induced Modification and Aggregation of Proteins and Hardening of Texture in a Protein Bar Model System . JAFC (accepted in press)
• Wang, Q, He, L., Labuza, T.P. and Ismail, B. (2012) Structural Characterization of Partially Glycosylated Whey Protein as influenced by pH and Heat using Surface-Enhanced Raman Spectroscopy J Agr. Food Chem. (accepted in press)
• Rao, Q., Klassen, A., and Labuza, T.P. (2012) Storage stability of Food Protein Hydrolysates. Critical Reviews in Food Science and Nutrition. (accepted in press)
• Lumor, S.E., Fredrickson, N., Ronningen, I., Deen, B., Smith, K., Diez-Gonzalez, F. and Labuza, T.P. (2012). Detection and Inactivation of Saxitoxin in Skim Milk. Int. Journal of Food Protection 75(6): 1113-1116. Lumor, S.E., Fredrickson, N., Ronningen, I., Deen, B., Smith, K., Diez-Gonzalez, F. and Labuza, T.P. (2012).. Comparison, in Food Matrices, of the Presence of Shiga Toxin 1 (Stx1) as Determined by an ELISA and a Biological Activity Assay. 2012 Intl. J. Food Protection 75(2): 1113-1116. DOI 10.4315/0362-028X.JFP-11-381
• Lili, H., Deen, B., Pagel, A., Diez-Gonzalez, F., and Labuza, T. P, (2012) Concentration, detection and discrimination of Bacillus anthrasis spores in orange juice using aptamer based surface enhanced Raman Spectroscopy. Analyst, DOI: 10.1039/c0xx00000x
• Roos, Y.H., Franks, F., Karel, M., Labuza, T.P., Levine, H., Mathlouthi, M., Reid, M. , Shalaev, E. and Slade, L. (2012) Comment on the melting and decomposition of sugars. J. Agric. Food Chem., 2012, 60 (41), pp 10359-10362. DOI: 10.1021/jf3002526
• . Hartel, R. and Labuza, T.P. (2012). Shelf Life of High-Moisture Confections: The Case of the Hardening Marshmallow. Manufacturing Confectioner. Accepted, in press.
• Abstracts Labuza, T.P. and Ismail, B., (2012) Changes in Stability of Bioactive Peptides through Glycation with Short Dextrans. Euro Foods Water July 1-3 Helsinki Finland. (Oral)
• Labuza, T.P. (2012) Accountability and the new US Food Safety Modernization Act (FSMA). Dept of Pub;lic Health/Chinese Inst. Of Food Technologists Food Saftey Forum April Beijing China (Oral)
• Rao, Q., Schoenfuss, T., and Labuza T.P. (2012) Effect of water activity on selected physicochemical properties of a commercial hydrolyzed hen egg white powder during storage at 45C. Institute of Food Technologists Annual Meeting June Las Vegas Nevada (Poster)
• Ismail, B, Rao, Q., and Labuza T.P. (2012) Aggregation in dry and intermediate-moisture food matrices containing protein hydrolysates Institute of Food Technologists Annual Meeting June Las Vegas Nevada (Poster)
• Fisher, M.C., Rao, Q. and Labuza, T.P. (2012). Effect of moisture content and humectants on hen egg protein bar quality loss during storage. Institute of Food Technologists Annual Meeting June Las Vegas Nevada (Poster)
• Wang, Q., He, L., Ismail, B. and Labuza, T.P. (2012) Structural Characterization of Partially Glycosylated Whey Protein using Surface-Enhanced Raman Spectroscopy and Mass Spectrometry. Institute of Food Technologists Annual Meeting June Las Vegas Nevada (Oral)
• Deen, B., Rodda,T., He, L., Diez-Gonzalez, F., and Labuza, T.P., (2012) Rapid Detection and Discrimination of Bacillus Species Using Immunomagnetic Separation Combined with Surface-Enhanced Raman Spectroscopy. Institute of Food Technologists Annual Meeting June Las Vegas Nevada (Poster)
• Lumor, S.E. and Labuza, T.P. (2012) Detection and Inactivation of Saxitoxin in Skim Milk. Institute of Food Technologists Annual Meeting June Las Vegas Nevada (Poster).
• Labuza, T.P., Rao, Q. and Ismail, B., (2012) Aggregation in dry and intermediate-moisture food matrices containing protein hydrolysates USDA NIFA PI Pre-IFT meeting June Las Vegas Nevada (Poster)
• He, L. and Labuza, T.P. (2012) Development of an IMS_SERS method for rapid detection of pesticides on food surfaces USDA NIFA PI Pre-IFT meeting June Las Vegas Nevada (Poster)
• Hou, Zhe, He, L, Fink, R., Labuza, T.P., Diez-Gonzalez, F., and Sadowsky, M.J. (2012) Analysis of Escherichia coli K12 and O157:H7 Interaction with Lettuce Roots Using Surface Enhanced Raman Spectroscopy. Am. Soc. Microbiology

Progress 01/01/11 to 12/31/11

Outputs
OUTPUTS: As a result of our prior USDA NRI and MDA Grants on whey protein aggregation, we were able to obtain a grant from the American Egg Board at the start of 2011. This was for work on the physical chemistry and stability by both disulfide bond aggregation and through Maillard reactions in dry egg proteins (Dry egg white, Dry egg yolk) as a function of moisture in model dry (1-10% db) and intermediate moisture systems (15-25% db). The outputs were 2 poster papers presented at the annual 2011 IFT meeting and the same 2 at the Midwest Dairy Center meeting in Roseville with 100+ in attendance. One paper on physical chemistry of dry egg white and its hydrolysates was published in Food Chemistry, which has the highest impact factor of food chemistry journals. In addition we published a book chapter on use of DSC (differential scanning calorimetry) for measuring protein denaturation in the Encyclopedia of Dairy Chemistry. This preliminary work led to us being funded by a USDA NIFA grant ($500K) for 2012-13 to study the physical chemistry of protein hydrolysates and methods to improve stability of bioactive peptides. In addition we obtained $5000 from the Mid West Dairy Center for preparing a monograph on peptides. This preliminary work led to funding for 2012-13 by a USDA NIFA grant ($500K) for developing a Surface Enhanced Raman Spectroscopy (SERS) assay for pesticides on produce. We also got a Diary Research Inc. grant ($130K) for 2012 to develop a rapid SERS method for dairy protein allergens. Outputs included 7 published papers in high impact factor journals including Chemical Science and the Analyst: journals of the Royal Chemical Society. In addition 2 posters related to DHS work were presented at the International Association for Food Protection (IAFP) annual meeting in Milwaukee. This work led to us being asked to present 2 webinars on SERS applications (6/3/11; 9/29/11) each with at least 100 participants. In addition we presented a poster on this to the Nano Technology Workshop (11/15/11) at the Univ. of Minnesota. PARTICIPANTS: 1. Dr. Christy Haynes is a McKnight Assoc. Professor in the Department of Chemistry at the U of M. She did her PhD at Northwestern Univ. in Chicago, on SERS under the guidance of Professor Van Duyne, one of the founders of the plasmon amplification effect. Her guidance on understanding the spectra evolved from different materials was critical to the project. 2. Dr Sri Sreevatsan is a Professor in the school of Veterinarian Medicine (UM). His research is on large animal virology using single stranded DNA designed to bind tightly to disease viruses instead of using antibodies. We met through NCFPD. A. Lamont is his grad student Keith Warriner is the technician. Dr. Veeregowda was a visiting scientist. 3. Dr. Francisco Diez is a Professor in the Food Science and Nutrition Department (U of M) who teaches/researches in the area of Food Safety and Microbiology. He and Dr Labuza have collaborated on research for about 10 yrs.. He has significant funding from the USDA for work on E. coli O157:H7 and on safety of organic foods. His BL-2 lab was a critical to our progress. 4. Dr. Lily He, did her PhD work on SERS at the Univ. of Missouri and came here funded as a post-doc (now a Research Associate) on a grant we had from NCFPD for developing rapid assays for biological toxins that could be used by terrorists. She developed the collaborations with the above faculty. .She was critical in getting the Raman instrument loaned by Thermo Fisher Inc. to the U of M. While at Missouri, Lily developed a rapid test for melamine using SERS. She has trained all the users of the instrument. 5. Dr. Qinchun Rao, did his PhD at Florida State Univ. in the area of identification of foreign proteins in meat or fish, i.e. economic adulteration. His quantitative skills led him to be the lab manager. He became mentor to Jean-Carlo Rocco, a student visiting scientist doing a thesis for a degree from the Univ. of Udine in Italy. Dr. Rao has a solid background in protein biochemistry/biophysics. 6. Dr.'s Tim Deschaines and T. Strother are principle scientists at the Thermo Fisher facilities in Madison Wisc, where they work on Raman applications. Their suggestions on methodology has been very useful and they were very instrumental in working out the loan and purchase of a Raman instrument. 7. Dr. Stephen Lumor is a native of Ghana and did his PhD on synthesis of non-trans fats at the Univ. of Georgia. He key role was in integrating the ELISA tests we were using in the inactivation studies of protein toxins like ricin with biochemical measures of toxic activity. In the summer of 2010 and 2011 he managed 6 undergraduate students who had to be trained well to work with very hazardous material. He did a marvelous job at helping them learn how to do basic science. These included several students from Food Science (Neal Fredrickson, Bronwyn Deen, and Ian Ronnigan), two from Chemical Engineering (Aaron Hutt and Tim Blaisius) and one from the Chemistry Dept. at St Scholastica (Ken Smith) in Duluth MN as well as one of my grad students (Tom Rodda). TARGET AUDIENCES: Other faculty in Food Science or Public Health around the world Those in charge of Food Security at each food processing or dietary supplement facility Public Health personnel Commercial food analytical Media PROJECT MODIFICATIONS: Addition of the work on SERS because that was deemed a national priority in Food Security

Impacts
Key 1st outcomes were (1) building a foundation knowledge on protein stability during storage; and (2) building base knowledge on combining immuno-magnetic separation with SERS to work on rapid methods of detection and quantification. The 2nd outcome which was the creation of two multidisciplinary teams which were successful in getting new major grants in both research areas. The 1st team for protein bio-physical chemistry showed that disulfide aggregation occurs at low water to intermediate moisture content, insolubilizing whey proteins which can be inhibited by cysteine. This finding was applied to dry egg albumin and dry egg yolk proteins. For both proteins, if any reducing sugars are present, then Maillard aggregation reactions predominate: this knowledge is extremely important to protein/ energy bar makers. The 2nd team developed methods to combine the capture of agents, pesticides or unwanted proteins with single stranded DNA aptamers attached to magnetic beads. When placed on a silver dendritic nano surface upon which a mono-chromatic laser beam is shown, this produces an amplified (10^6) Raman fingerprint signal because of the plasmon effect generated by the nano surface. Levels of detection of ricin or egg albumin in milk are 5 nanogram/mL. These 2 teams responded to requests for proposals which then lead to the 3rd outcome, i.e. success in obtaining the following: 1. USDA NIFA grant ($500K) on bio-physical chemistry /stability of bioactive peptides and their dextran adducts (2012-2013); 2. USDA NIFA Grant ($500K) to study nano-technology based methods for rapid detection of both legal and illegal pesticides on the surface of fresh produce as well as on the surface of food processing equipment; 3. A $126,000 grant from Dairy Research Inc. for a rapid test for milk protein allergens using the IMS-SERS; 4. A $5000 grant from the Minn. Dairy Assoc. for a monograph on stability of peptides; 5. Extension of the AEB grant ($55K) for 6 months; and 6. A $256K grant from the Dept of Homeland Security for a similar test for anthrasis spores.

Publications

• Meeting Abstracts and or proceedings from oral or poster presentations. ( Total poster presentation 8 Oral Presentations 6). 2011.
• Labuza, T.P. 2011. Accountability and the New Food Safety Modernization Act. Program Book: Risk Communication and Food safety Management for the Whole Food Supply., pp 44-46. IUFOST and CIFST publication. 4/21-2/2011, Bejing China (oral)
• Labuza, T.P. 2011. Water Relations in Foods: Paradigm shifts to supplant cook and look. in Food Process Engineering in a changing World. 5/22-25/2011. Athens, Greece. iCEF 11 Congress Proceedings. Vol. 1:pp 75. Editors P. Taoukis, N. Stoforos, V. Karathanos and G. Saravacos. Cosmoware Ag, Athens Greece (oral)
• He, L., and Labuza T.P. 2011. Aptamer conjugated dendritic silver nano-structure and its application in SERS detection of a protein toxins. 7th Annual Minnesota Nanotechnology Workshop. 11/15/11. Minneapolis, MN. (poster presentation)
• Refereed Research Papers (total 9 published, 3 in press) 2011
• Lumor, Stephen E. , Diez-Gonzalez, F., and Labuza, T.P. 2011. Detection of Warfare Agents in Liquid Foods Using the Brine Shrimp Lethality Assay Jr. Food Science 76:T16-T19; DOI 10.1111/j.1750-3841.2010.01966.x.
• Lumor, Stephen E, Hutt A., Ronnigen, I., Diez-Gonzalez, F., and Labuza, T.P. 2011. Validation of Immunodetection (ELISA) of Ricin Using a Biological Activity Assay. Jr. Food Science 76(1) C112-116 DOI 10.1111/j.1750-3841.2010.01943.x
• He, L., Lamont, A., Veeregowda, B., Sreevatann, S., Haynes, C.L., Diez-Gonzalez, F., and Labuza, T.P. 2011. Aptamer based surface-enhanced Raman scattering detection of ricin in liquid foods. Chemical Science 2:1579-1582. DOI: 10.1039/C1SC00201E
• He, L., Haynes, C.L., Diez-Gonzalez, F., and Labuza, T.P. 2011 Rapid Detection of a Protein Toxin Surrogate in Milk using IMS-SERS. Jr. Raman Spectroscopy 42: 1428-1434. DOI 10.1002/jrs.2880
• He, L., Rodda, T., Haynes, C.L., Deschaines, T., Strother, T., Diez-Gonzalez, F., and Labuza., T.P. 2011. Detection of a Foreign Protein in Milk Using Surface-Enhanced Raman Spectroscopy Coupled with Antibody-Modified Silver Dendrites. Analytical Chemistry 83:1510-1513 .DOI dx.doi.org/10.0121/ac1032353
• He, L., Haynes, C.L., Deen, B., Rodda,T., Ronnigan, I., Blasius, T., and Labuza, T.P. 2011. Rapid Detection of Ricin in Milk using Immunomagnetic Separation Combined with Surface-Enhanced Raman Spectroscopy; J. Food Science 76 (5): N49-N53
• Lamont, E.L., He, L., Warriner, K., Labuza, T.P., and Sreevatsan, S. 2011. A single DNA aptamer functions as a biosensor for ricin. 2011 Analyst 136: 3884-389. DOI: 10.1039/c1an15352
• Rao, Q., and Labuza , T.P. 2011. Effect of moisture content on selected physicochemical properties of two commercial hen egg white powders. Food Chemistry doi:10.1016/j.foodchem.2011.10.10
• Liu, D., Zhou, P., Liu, X., and Labuza, T.P. 2011. Moisture-Induced Aggregation of Alpha-Lactalbumin: Effects of Temperature, Cations, and pH. Jr. Food Science 76(6):C817-823 DOI 10.1111/j.1750-3841.2011.02277.x
• Lumor, S.E., Fredrickson, N.R., Deen, B.D., Ronningen, I., Smith, K. F. Diez-Gonzalez and T.P. Labuza. 2011. Comparison between ELISA determination of Shiga toxin 1 (Stx1) in food matrices with a biological activity assay. Journal of Food Protection (in press)
• Lumor, S.E., Deen, B., Ronningen, I., Fredrickson, N., Smith, K., Diez-Gonzalez, F., and Labuza, T.P. 2011. Assessment of the potential inhibition of ricin toxicity by lactose in milk. Jr. Food Science (in Press)
• Lumor, S., Deen, B.D., Ronningen, Smith, K., Fredrickson, N.R., Diez-Gonzalez, F., and Labuza, T.P. 2011. Detection and inactivation of saxitoxin in skim milk. Journal of Food Protection (in press)
• Book Chapters.(Total 1) 2011.
• Zhou, P. and Labuza, T.P. 2011. Differential Scanning Calorimetry in Encyclopedia of Dairy Sciences. , pp 256 to 263., Editors: J.Fuquay, P. McSweeney and P. Fox. Academic Pres; San Diego
• Labuza, T.P. 2011. Prof. Marcus Karel: Building Paradigms for Food Engineering and Material Science as influenced by Water. In Food Process Engineering in a changing World. 5/22-25/2011. Athens, Greece. iCEF 11 Congress Proceedings. Vol. 1:pp 755-757. Editors P. Taoukis, N. Stoforos, V. Karathanos and G. Saravacos. Cosmoware Ag, Athens Greece (oral)
• Labuza, T.P. Diez-Gonzalez,F., Lumor,S.E, Deen,B.D., Ronningen,I. and Smith, K. 2011. Abstract # P1-42 Effect of Lactose on the Biological Activity of Ricin. 100th Annual meeting of the International Association for Food Protection. Milwauke, WI 8/1-4/2011. (poster presentation)
• Viazis, S., Labuza, T.P., and Diez-Gonzalez, F., 2011. Abstract # P1-78. Inactivation Kinetics of Escherichia coli O157:H7 on Hard Surfaces by Use of a Bacteriophage Mixture 100th Annual meeting of the International Association for Food Protection. Milwauke, WI 8/1-4/2011. (poster presentation)
• Qinchun, R., and Labuza, T.P. 2011. Abstract # 288-11. Effect of moisture content on glass transition and protein aggregation of egg white powder Annual meeting of the Institute of Food Technologists. New Orleans , LA . 6/11-14/2011. (poster presentation)
• Fisher, C., and Labuza, T.P. 2011. Abstract # 236-58. Effect of storage temperature on the quality losses for model egg protein bar system. Annual meeting of the Institute of Food Technologists. New Orleans , LA . 6/11-14/2011. (poster presentation)
• He, L.,, Haynes, C.L., Diez-Gonzalez, F., and Labuza, T.P. 2011. Abstract # 161-04. Rapid detection of a foreign allergenic protein in milk using surface-enhanced Raman Spectroscopy coupled with antibody-modified silver dendrites Annual meeting of the Institute of Food Technologists. New Orleans , LA . 6/11-14/2011. (poster presentation)
• Lumor, S.E., Deen, B., Ronningen, I., Fredrickson, N., Smith, K., Diez-Gonzalez, F., and Labuza, T.P. 2011. Assessment of the potential inhibition of ricin toxicity by lactose in milk. Midwest Dairy Center Annual Meeting 7/17/11 (poster presentation).
• Labuza, T.P. 2001. The use of IMS-SERS for rapid detection of protein allergens in dairy products. Midwest Dairy Center Annual Meeting Proceedings 7/18/11 (oral presentation)
• Lumor, S.E., Deen, B., Ronningen, I., Fredrickson, N., Smith, K., Diez-Gonzalez, F., and Labuza, T.P. 2011. Assessment of the potential inhibition of ricin toxicity by lactose in milk. National Center for Food Protection and Defense Themes Meeting 9/26-27/11 Minneapolis UM McNamarra Center (poster presentation)
• He, L., and Labuza T.P. 2011. Aptamer conjugated dendritic silver nano-structure and its application in SERS detection of a protein toxins. National Center for Food Protection and Defense Themes Meeting 9/26-27/11 Minneapolis UM McNamarra Center (poster presentation)
• Labuza, T.P. 2011. The physical chemistry of reducing salt in foods. Food Science and Nutrition Advisory Committee. 10/12/2011 St. Paul, MN. (oral)

Progress 01/01/10 to 12/31/10

Outputs
OUTPUTS: The 1st project focuses on investigating the ability of polyols and cysteine to improve the stability of whey proteins in an intermediate-moisture protein/energy bar model system. The stability problem is with respect to both hardening of these whey protein-based bars due to Maillard browning and di-sulfide interactions and loss of available lysine (nutritional impact) during storage. Prior work showed that maltitol should be successful in replacing HFCS so this year's focus was an accelerated shelf life test as well as the ability of cysteine to inhibit hardening. This work started in 2002 as a Midwest Dairy Center project (Check-off $), that morphed into a USDA NRI 3 year grant and beginning this year, a 2 year grant from the American Egg Board (Check-off $). It has funded 4 post docs and 3 MS degrees and supported 6 undergrads. Three post docs got faculty or research appointments at Jiangnan Univ and Univ. of Wisconsin where they continue this work. One, Zhou, was selected as the outstanding young research food scientist in the world by IUFOST this year. An MS grad was hired by the Irish Dairy Board to continue this at Univ. College Cork, Ireland. It has produced 8 refereed publications with 3 more submitted and 2 in preparation, 7 of them are in the Jr. Agric. Food Chem, the top refereed research journal for food chemistry with widespread reading. The results of this work in 2010 have been presented 1) nationally at the IFT (3 venues) including the Jiangnan Univ. US Alumni Assoc.; and 2) internationally to an international meeting on water in foods held in Germany and at the IUFOST 15th bi-annual meeting in Cape Town. In each case there were approximately 200 or more professionals attending. A 2nd project supported on USDA check-off money from the Almond Board of California deals with moisture properties of almonds in different styles, e.g. whole vs. slivered. Isotherms and moisture diffusion coefficients were determined for 58 different varieties/styles and this year the results were presented to over 250 growers/processors. This work will allow users to estimate the potential for loss of almond crunchiness in different packages and in different venues like trail mixes. Related to this, 5 industrial short courses were held relating temperature and water activity to shelf life and an IFT Webinar was presented with 300 attendees. A new 3rd area added in this year and funded by US Dept. Homeland security is the identification of allergens at the nano level in food stuffs using IMS-SERS (Immuno Magnetic Separation-Surface Enhanced Raman Spectroscopy). Initial work suggests it is feasible, enough to get the loan of a state of the art Raman instrument from Thermo Fischer scientific. Our findings suggest detection an the 0.2 micrograms per liter level. PARTICIPANTS: Assoc. prof. Peng Zhou, State Key Laboratory for Food Science, Jiangnan Univ. Wuxi PRC; Asst. Prof. Xiaoming Liu, School of Food Sc. & Tech., Jiangnan U, Wuxi, PRC; Dr. Michael Uzzan, Technion University, Haifa, Israel; Qinchun Rao, Post-Doc, U of MN; U of MN MS Graduates: Amy Tran, University College, Cork, Cork, Ireland; Heather Taterka, Fulbright Scholarship student posted MS work at the Norwegian National Research Center in Trondheim, Norway; Na Wang, Schroeder Cheese Milw., WI; U of MN food science undergraduate students: Sarah Gutkowski, Rachel Kyllo, Brian Folger, Kristen Ruuf, Ian Ronnigan and Neal Fredrickson. Engineering undergraduate students Aaron Hutt, Cherry Lam, Katlin Sautner, Tim Blaisius and Brian Blonigan, Chem. undergraduate, St. Scholastica, Duluth, MN Kenny Smith. TARGET AUDIENCES: The food industry; most of the participants at the various presentations would be food product development scientists; Twelve undergraduate students over the past 6 years who want to do more science beyond a BS degree. PROJECT MODIFICATIONS: We had added a new area with respect to detecting allergen protein in complex food maticies using a new technique of IMS-Seres. Detection is within 20 to 30 min at the .2 ppb level. We are seeking funding for this.

Impacts
Impact 1. This research was prompted by the findings in pharmaceutics paper on protein aggregation in the dry and semi-moist state of a dry inhalation insulin drug. This drug was designed for diabetics and removed the need to inject into the blood stream. Unfortunately the insulin became insoluble and caused granulomas to form in the lungs and the drug was recalled by the FDA. In 2001, our lab was consulted by a number of protein bar manufacturers to help them find out the cause of hardening without moisture loss. We had been working on this on another project related to cookie hardening and postulated 3 modes: (1)disulfide aggregation; (2) Maillard browning aggregation since most used high fructose corn syrup (HFCS) and (3) HFCS crystallization leading to loss of plasticizer. In testing these modes, the 1st significant outcome was that replacing HFCS with a polyol like maltitol eliminates browning and makes the bar softer for at least one year and maintains protein quality. Secondly, replacing the whey isolate by 25% with a medium degree (5 to 7%) hydrolysate lowers the glass transition temperature, also making the bar softer. And thirdly, adding an optimum amount of cysteine inhibits the di-sulfide aggregation and preserves protein quality. Thus a combination of all 3 approaches can be used by protein/energy bar makers to get an acceptable shelf life and ensure consumers that the bar actually delivers high quality protein. Thus there is a change in both knowledge and actions, especially by the manufacturers. This research led to the American Egg Board to fund us in this area as the use of egg proteins has been low because of it's tendency to harden due to the high -SH group content. Impact 2. We have found that using an accelerated test for shelf life of protein bars by storing for one month at 45 C, translates into ~ 1 year life at 23 C. This will significantly shorten product development time. Impact 3. Our work on almonds has led to 3 potential solutions to problems facing the industry. The first is that most producers are selling product at moistures much higher than the BET monolayer value. This practice allows them to sell more water as almonds but shortens shelf life in terms of the moisture gain to transition from a crunchy to a mealy unacceptable product. Having the isotherm will allow them to better target finished moisture. Impact 4. California being the biggest "country" for almond production also allows them to be the biggest exporter at over 1 billion pounds per year. However, a lot of almonds that reach China, the biggest buyer, are rejected upon delivery because on either browning or mold growth. The product is shipped in metric ton size with a poor water vapor barrier. Our diffusion data shows that the surfaces reach outside humidity very quickly leading to moldiness at >75% RH, conditions the product faces in the 3 weeks shipboard transit to Shanghai. A 6 mil polyethylene overwrap should give enough protection.

Publications

• Leishman, O., Labuza, T.P. and Gonzalez-Diez, F. 2010. Hydrophobic Properties and Extraction of Bacillus anthracis Spores from Liquid Foods J. Food Microbiology, 27:661-666. http://dx.doi.org/10.1016/j.fm.2010.03.001
• Leishman, O.N., Johnson, M.J., Labuza, T.P. and Diez-Gonzalez, F. 2010. Survival of Bacillus anthracis Spores in Fruit Juices and Wine, J. Food Protection, 73(9):1694-1697.
• Bott, R.F., Labuza, T.P. and Oliveira, W.P. 2010. Stability testing of spray and spouted bed dried extracts of Passiflora alata. Drying Research & Technology, 28:1255-1265.
• Mortenson, M.A., Labuza, T.P. and Reineccius, G.A. 2010. Moisture sorption isotherms for un-modified and OSAn-substituted dextrin and gum acacia used as carrier materials for spray dried encapsulation of flavoring materials. Intl. Journal of Food Properties, Vol 13 (5):992-1001. http://DOI:10.1080/10942910902930643
• Zhou, P. and Labuza, T.P. Moisture-introduced aggregation of whey proteins, 2009. International Journal of Science & Marketing for Nutraceutical Actives Raw Materials Finished Products, 8(4):29-32. (paper not published until April 2010 with 2009 date
• Zhu, D. and Labuza T.P. 2010. Effect of cysteine on protein bar hardening in WPI/buffer model systems and the elucidation of the underling mechanism. J. Agric. Food Chem., 58: 7970-7979 http://DOI:10.1021/jf100743z
• Floros, J.D., Newsome, R. Fisher, W., Barbosa-C'anovas, G.V., Chen, H., Dunne, C.P., German, J.B., Hall, R.L., Heldman, D.R., Karwe, M.V., Knabel, S.J., Labuza, T.P., Lund, D.B., Newell-McGloughlin, M., Robinson, J.L., Sebranek, J.G., Shewfelt, R.L., Tracy, W.F., Weaver, C.M. and Ziegler, G.R. 2010. Feeding the World Today and Tomorrow: The importance of Food Science and Technology; Comprehensive Reviews in Food Science and Food Safety, Vol. 9 (9): 572-599 http://onlinelibrary.wiley.com/doi/10.1111/j.1541-4337.2010.00127.x/p df
• Labuza, T.P., Labuza, T.J., Labuza, P.S. and Labuza, K.M. 2010. Soft Condensed Matter: A perspective on the physics of food states & stability in Water Properties in Food, Health, Pharmaceutical and Biological Systems, Chap. 7, in ISOPOW 10. D. Reid, T. Sajaanantakul, P. Lilford and S. Charoenrein (Ed.), Willey-Blackwell Press, Ames, IA, p. 87-114. ISBN-13: 978-0-8138-1273-1
• Bott, R.F., Souza, C.R.F., Labuza, T.P. and Oliveira, W.P. 2010. Estabilidade de extratos secos de passiflora alata obtidos por spray drying e leito de jorro. Proceedings of the 24th Brazilian Congress of System Particulates

Progress 01/01/09 to 12/31/09

Outputs
OUTPUTS: The first project focused on determining the factors influencing the stability creatine in a liquid delivery system. It is widely used as a dietary supplement for athletes and body builders. One of the main obstacles in creatine usage as a dietary supplement is the fact that creatine is a weak base that is easily degraded to creatinine, a nonactive metabolite, especially in acid environments. A second project investigated the influence of polyols on the stability of whey proteins in an intermediate-moisture food model system and to elucidate the effect of polyols on the hardening of whey protein-based bars during storage. The objective was to determine if the use of these polyols would improve the shelf-life of whey protein-based energy bars. Thus, the effect of adding four major polyols (glycerol, propylene glycol, maltitol and sorbitol) to model bar systems containing whey protein isolate, polyols, and water was investigated. The third project focused on a food safety issue. The primary objective was to estimate the time to detect (TTD) Listeria monocytogenes in frankfurters from inoculum levels below the detection limit, and model the TTD at three temperatures to deduce a safety-based shelf life equation. In this work L. monocytogenes was inoculated on frankfurters at four inoculum levels and was then held at 4, 8, and 12C until the time L. monocytogenes populations reached a detectable limit of at least 2 CFU/g. Based on the TTD from each inoculum level and at each temperature, a shelf life model was constructed to predict the detection or risk levels reached by L. monocytogenes on frankfurters. The results of these projects have been presented 1) locally to The Food Industry Center annual symposium, U of M Coffman Union, Fairview South Medical Center, Nutrition Grand Rounds; 2) nationally to the National Grocers Assoc. annual meeting, Las Vegas Nevada; Conference on Food Engineering (COFE) in Columbus Ohio, the annual meeting of USDA CREES PIs NRI grants in New Orleans, IFT annual meeting in New Orleans; and to the Assoc. of Food Protection in Washington DC; and 3) internationally to WHO in Geneva, Switzerland; TNO Smart Packaging Workshop in NXP Holst Center Eindhoven, Netherlands as well as via the www (e.g., Traceability IFT Webinar, Designer and Presentation). PARTICIPANTS: Peng Zhou, School of Food Sc. & Tech., Jiangnan U, Wuxi, China Francisco Diez-Gonzalez, U of M Dept. of Food Science and Nutrition Michael Uzzan, Technion University, Haifa, Israel Amy Tran, University College, Cork, Cork, Ireland Amit Pan, Campbell Soup Co., Camden, NJ TARGET AUDIENCES: Nothing significant to report during this reporting period. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

Impacts
Creatine stability: Creatine degradation to creatinine (no biological activity) in combinations of glycerol and pH 4.0 buffer solutions followed first-order kinetics up to a point where degradation started to level off, generally beyond the first half-life. Practical data were reported for a wide range of water activity (aw) values (0.31-0.983) at 4 C, 23 C, and 35 C. Creatine degradation did not exhibit a dilution effect, that is a decrease in rate above an aw of 0.7, as is found for both microbiological growth and chemical reactions in semisolid food matrix systems. The temperature dependence obeyed the Arrhenius relationship with an energy of activation of about 20 kcal/mol at aw &#8805;&#61472;0.68 increasing to 23 kcal/mole below that aw. In addition, a semilog plot of half-life as a function of aw at each temperature follows a predicted straight line. The pH and assumed liquid viscosity increase through increased glycerol concentration were not able to completely explain the decrease in rate of degradation as a function of aw. Furthermore, we confirmed that creatine stability in the crystal form is very high as long as it does not reach the deliquescence state. Hardening of whey protein-based bars: The results of this study showed that glycerol was the most effective polyol in lowering water activity and providing a soft texture of intermediate-moisture foods, followed by sorbitol and maltitol. These three polyols stabilized the native structure of whey proteins, provided a desired texture, and slowed the hardening of the protein bar systems. Propylene glycol should not be used in whey protein-based high protein intermediate-moisture foods because it caused irreversible protein aggregation as observed by differential scanning calorimeter and Fourier transform infrared spectroscopy. The aggregation of the whey proteins caused hardening of the bar texture during storage, causing loss in product quality. Listeria monocytogenes: The time to detection (TTD) increased with reductions in inoculum size and storage temperature. At 4C the TTDs (standard error) observed were 42.0 +/- minus 1.0, 43.5 +/- 0.5, 50.7 +/-1.5, and 55.0 +/- 3.0 days when the inoculum sizes were 0.1, 0.04, 0.01, and 0.007 CFU/g, respectively. From the same corresponding inoculum sizes, the TTDs at 8C were 4.5 +/- 0.5, 6.5 +/- 0.5, 7.0 +/- 1.0, and 8.5 +/- 0.5 days. Significant differences (P < 0.05) between TTDs were observed only when the inoculum sizes differed by at least 2 log. On a shelf life plot of ln(TTD) versus temperature, the Q10 (increase in TTD for a 10C increase in temperature) values ranged from 24.5 to 44.7 and with no significant influence from the inoculum densities. When the observed TTDs were compared with the expected detection times based on the data obtained from a study with an inoculum size of 10 to 20 CFU/g, significant deviations were noted at lower inoculum levels. These results can be valuable in designing a safety-based shelf life model for frankfurters and in performing quantitative risk assessment of listeriosis at low and practical contamination levels.

Publications

• Uzzan, M., Necherbecki, J., Zhou, P. and Labuza, T.P. 2009. Effect of water activity and temperature on the stability of creatine during storage. Drug Development and Industrial Pharmacy, 35(8):1003-1008.
• Xiaoming L., Zhou, P., Tran, A. and Labuza, T. P. 2009. Effects of polyols on stability of whey proteins in intermediate moisture model systems. Journal of Agricultural Food Chemistry, 57(6): 2339-2345.
• Pal, A., Labuza, T.P. and Diez-Gonzalez, F. 2009. Time-to-detect modeling of L. monocytogenes growth on frankfurters with initial inoculum below standard detection limit. Journal of Food Protection, 72(2):1878-1884.
• Zhou, P. and Labuza, T.P. 2009. Moisture-introduced aggregation of whey proteins. International Journal of Science & Marketing for Nutraceutical Actives Raw Materials Finished Products 8(4).
• Li, T.C., Zhou, P., Labuza, T.P. 2009. Effects of sucrose crystallization and moisture migration on the structural changes of a coated intermediate moisture food. Frontiers of Chemical Engineering in China 3(4): 346-350.
• Yildiz, F., Labuza, T.P. and Besri, M. Advances in Food Biochemistry. 2009. Ed. Yildiz, CPC Press - Taylor Francis Group, Boca Raton, FL. 800 pp.

Progress 01/01/08 to 12/31/08

Outputs
OUTPUTS: The research includes: (1) loss of protein quality and hardening of protein based bars supported by a USDA NRI grant; (2) evaluation of a new automated sorption device supported by Decagon Corp; (3) determination of the kinetics of inactivation of bioterror agents by heat and biocides; (4) development of rapid methods to detect anthrax spores in milk and juice systems; area (5) kinetics of growth of Listeria in Ready to Eat meats; and (6) state diagrams and crispness of foods. Area 1: Early work showed that whey protein based bars hardened much faster that did soy based bars. At the end of last year we also got funding from Davisco, a producer of the whey proteins we are using. The work includes: (1) the basic chemistry of the hardening process and (2) practical solutions to slow hardening. Three mechanisms occur: (1) simple water diffusion into the protein particles causing loss of plasticizer volume between the particles; (2) inter-protein disulfide bond formation; and (3) protein interactions that occur through the Maillard browning reaction which not only makes them hard but also causes loss of protein quality. An additional finding was that there was also a direct action of a commonly used humectant, propylene glycol (PG), with the whey protein causing it to aggregate and become totally insoluble. Several years ago the FDA banned the use of PG in intermediate moisture pet foods as it caused an interaction with the hemoglobin in the blood cells of cats, (Heinz body formation) causing severe anemia. The results of these studies were presented at 2 dairy protein advisory committee meetings at the DMI headquarters in Chicago, at the international Dairy Meeting San Francisco (> 300 attendees), at the International Whey Conference (invited) in Paris (> 250 attendees), a poster paper and an oral invited presentation for the Dairy Division of IFT at the IFT Annual meeting (> 200 attendees), along with a poster at the USDA NRI PI meeting in New Orleans, a poster at the IUFOST meeting in Shanghi China, and a invited oral presentation at the Euro Foods Water Congress in Stuttgart GR ((200 attendees). We also transferred our results to the Am. Dairy Export Council (ADEC) who are making protein bars to help stave off malnutrition in Ethiopia. We have also presented at Davisco, a sponsor of the research. In area 2 we have finished the evaluation of a new instrument for automated isotherm determination (Decagon) in comparison to the standard static method and an automated instrument used in the pharmaceutics area. We have found the instrument to be acceptable. In area 3 under a DHS grant, we have completed work on anthrax spores and ricin indicating that heat alone using the typical dairy processing equipment is not sufficient and there is a need for using heat with biocides. Several talks have been made at dairy companies. In area 4 under another DHS grant we have evaluated a centrifugation/phase specific method which has promise. In area 5 we have made presentations at several meat processors, at the Int. Food Protection Assoc. meeting and at the 3rd Cold Chain meeting in Germany. In Area 6 we have presented at 3 international meetings. PARTICIPANTS: Protein area: Dr. Peng Zhou (former post-doc and PhD from Cornell Univ. is now a Professor at Jiangnan Univ. (Wuxi, PRC) in the State Laboratory in the Food Science and Technology Dept. Lloyd Metzger's former post doc Xiaoming Lui (Phd Wash. State U) is also a professor at Jiangnan Univ. PRC. Zhou trained students in use of the DSC, TAXT, and FTIR. Lui worked on protein aggregation during thermal processing. Current Post Doc Dan Zhu, prior Post-docs in Japan and the Dairy Research Center, U. of WI. Amy Tran, MS student will graduate in February, did work on protein nutritional quality loss in the bars. Heather Taterka, MS student who is evaluating the influence of different humectants on quality changes during storage. She will become a Fulbright Fellowship for a position in Norway. Tim Blasius (Jr. in Chem. Eng.) did an Undergraduate Research Opportunities Program (UROP) study in our labs during spring 2008 and continues as a lab helper. He will have a summer internship at Malt O- Meal next summer. Katlin Sautner (Soph. Chem. Eng) is a lab assistant along with Jackie Koch (Sr. Food Sci.) who is doing an honors thesis in this area. Laurie and Matt Davis of Davisco who have supported this work with a donation of whey proteins and with a substantial monetary gift. Other areas: Bioterror Research (DHS grant) Sa Xu (post-doc) came to us from Iowa State where she was doing a post-doc. Sa developed the specifics for study of the kinetics of inactivation of bioterror agents. She left after 2 years to head up microbiology at Land O' Lakes in Minnesota. Stephen Lamour, from Ghana who finished his PhD at the University of Georgia in food chemistry and is replacing Sa Xu. Na Wang, a student from China with a degree in Food Engineering is working on inactivation kinetics of ricin and shigatoxin with the help of Tim Blasius and Jackie Koch, a senior in Food Science who also did a UROP project last year. Her work this year will count towards the thesis requirement in the College Honors program. Oriana (Rabbe) Leishman is close to finishing her PhD on rapid methods for antharx detection in fluid foods. She came to us after finishing an MPH degree at Minnesota. The Minn. Dept. of Ag. (H. Kassenborg, supported the UROP project of Ann Wiley (Nutrition) and the MS/MPH thesis work of Peri Periakruppan and Ryan Newkirk on development of a model for the milk flow chain in MN. Aflatoxin Project (grant from Thermo-Electron: Corinne Otte, an MS student who finished her degree in June, helped by Sorcha Kennedy, an exchange student from the Univ. of Cork Ireland. They developed the SOPs needed to work with aerosilized toxin. Gerry Broski, who heads the Food Safety group in Thermo-Electron's Process Analytical Technology Division (Blaine MN), Moisture Sorption Work: Jody Shebuski-Shands who will finish her MS in February after obtaining 20 years experience in the food industry including stints at Pillsbury, General Foods, Kraft and General Mills. She had help from Tim Blasius and Brian Blonigen, (Sr. Chem. Eng.) who also did a UROP project. Listeria kinetics: Amit Pal who finished his PhD this year and in now at Campbell Soup and P. Minerich, VP R&D Hormel. TARGET AUDIENCES: Our audiences are food scientists interested in (1) the chemistry of protein quality change, those in protein industry interested in selling quality proteins and the nutritional bar manufacturers (a $1 billion market; (2) the cereal industry who needs means to detect and remove aflatoxin in the grain delivery chain; (3) those in the management and regulation of the food chain with respect to protecting and defending the food supply in a bioterror event; (4) those who work with dry and intermediate moisture foods wishing to find better method for developing sorption isotherm and (5) the processed RTE meat industry. PROJECT MODIFICATIONS: As seen we have branched out in several new areas because of the funding we received.

Impacts
In area 1 (protein bars) we have given talks or poster presentations at 2 international meetings in the US and 3 foreign countries. We have sent information to at least 20 companies who have heard the presentations, 3 of whom are the major manufacturers of whey proteins. They are translating the findings to their clients who are making protein bars. Our work with ADEC will insure that the children getting the protein bars will be of high enough quality to be consumed, ie not hard and of high nutritional value. Because of funding other than the USDA we were able to travel outside the USA to present to a more global audience. The response at the whey conference in Paris was exceptionally good as many foreign companies that are involved with bars were present and requested further material. A recent visit to the grocery store shows that one of our findings, i.e. the negative reaction of PG with whey protein, may have lead to many bar makers switching over to maltitol despite it's high cost. In area 2 we have just finished evaluation of the instrument and thus have only presented internally in general seminars. A poster paper has been accepted for by the 2009 Annual meeting. In area 3 and 4 which is funded by the Dept. of Homeland Security (DHS) through the National Center for Food Protection and Defense (NCFPD) we have presented at Center meetings and 3 international meetings in the past 3 years. As the material is sensitive our presentations have been limited.

Publications

• Raabe, O.N., Labuza, T.P., and Diez-Gonzalez, F . 2008. Distribution and concentration of bacillus anthracis spores added to raw milk following simple and continuous centrifugation. Abstrct book, NCFD Annual Meeting Chaska Mn
• Labuza, T., Pal, A., and Diez-Gonzales, F. 2008. Time to detect for safety-based shelf life dating of RTE foods. 3rd Annual Cold Chain Symposium, Bonn ,Germany
• Abstracts (2008) Labuza,T.P., Payne, C., and Labuza, T.J. 2008. Relationship between the state diagram based on brittle ductile transition and the sensory crispness of a hard cracker. Proceedings of the Crispy Crack Symposium Univ. Wageningen, Wageningen, Netherlands.
• Labuza, T. P. , Shands, J., and Zhou, P. 2008. Antiplasticizing and hardening of crisp cereals. Abstract Book 5th Euro Foods Water Conference, Stuttgart, Germany
• Kang, J. and Labuza, T.P. 2008. Glass Transition Temperatures of Sucrose-Water and Trehalose-Water Systems: A Population Modeling Approach. Abstract book Am. Assoc. Pharmaceutical Science, San Diego, CA.
• Technical Article. Isengard, H., Lillford, P., Reid, D. and Labuza, T.P. 2008. Taking the measure of water. IUFOST http://www.iufost.org/reports_resources/bulletins/documents/IUF.SIB.W aterinfood_000.pdf
• Newkirk, R., Bender, J., and Labuza, T.P. 2008. Modeling of the milk distribution system for a bioterror attack. Abstract book NCFD Annual Meeting Chaska
• Otte, C., Kennedy, S. and Labuza, T.P. 2008. Front faced fluorescence spectroscopy in process analytical technology for detection of aflatoxin. Abstract book IFT Annual meeting, New Orleans, LA.
• Raabe, O.N., Labuza, T.P., and Diez-Gonzalez, F. 2008. Development of a PCR positive control protocol for detecting the presence of bacillus anthracis plasmid pXO2. Abstract book Am. Microbiology Society Annual Meeting, San Francisco CA
• Raabe, O.N., Labuza, T.P., and Diez-Gonzalez, F. 2008. Development of a PCR positive control protocol for detecting the presence of bacillus anthracis plasmid pXO2. Abstract book NCFD Annual Meeting Chaska, Mn
• Pal, A., Labuza, T.P. and Diez-Gonzalez. F. 2008. Evaluating the growth of Listeria monocytogenes in refrigerated ready-to-eat frankfurters, Influence of strain temperature, packaging, lactate and diacetate, and background microflora. Intr. Jr. Food Protection 71(9):1806-1816
• Pal, A., Labuza, T.P. and Diez-Gonzalez. F. 2008. Shelf Life Evaluation for Ready-to-Eat Sliced Uncured Turkey Breast and Cured Ham under Probable Storage Conditions based on Listeria monocytogenes and psychrotrops. International Jr. Food Microbiology 126:49-56
• Conference Proceedings: Labuza, T.P., Zhou, P., Liu, X., Davis,L., and Tran, A. 2008 Protein aggregation and hardening of whey based protein bars during storage. 5th International Whey Conference Paris, France Proceedings http://www.iwc-2008.org/home.asp
• Refereed Journal Articles: Zhou, P., Liu, X., and Labuza, T.P. 2008. Effects of moisture-induced whey protein aggregation on protein conformation, the state of water molecules, and the microstructure and texture of high-protein-containing matrix, Journal of Agricultural and Food Chemistry, 56: 4534-4540
• Zhou, P., Liu, X. and Labuza, T.P. 2008. Moisture induced aggregation of whey proteins in a protein- buffer model system. Journal of Agricultural and Food Chemistry, 56: 2048-2054
• Xu,S., Labuza, T.P., and Diez-Gonzalez, F. 2008. Inactivation of bacillus anthracis spores by a combination of biocides and heating at high-temperature short-time (HTST) Pasteurization Temperatures. Jr. Appl.& Environ Micro. 74(11):3336-3341
• Pal, A., Labuza, T.P. and Diez-Gonzalez. F. 2008. Comparison of Primary Predictive Models to Study the Growth of Listeria monocytogenes at Low-Temperatures in Liquid Cultures and Selection of Fastest Growing Ribotypes in Meat and Turkey Product Slurries, J. Food Microbiology 25:460-470
• Labuza, T.P., Pal, A. Belina, D and Diez, F. 2008. Time to detect for safety based shelf life of RTE foods. Proceedings of the 3rd Chilled Foods Summit. Pg 55-56 Judith Kreyenschmidt Editor Univ. Bonn, Germany
• Abstracts: Labuza, T.P. 2008. Influence of moisture on aggregation of protein during storage. 5th Euro Foods Water Conference, Stuttgart, Germany.
• Labuza, T.P., Tran, A., Taterka, H., and Zhou P. 2008. Mechanisms for Whey Protein aggregation during storage. Abstract Book of the 5th International Whey Congress (IWC), Paris, France.
• Zhou, P., Liu, X., Tran, A., and Labuza, T.P. 2008. Aggregation of whey protein in nutritional bars. Abstract Book IFT Dairy division for IFT Annual Meeting, New Orleans, LA.
• Labuza, T., Zhu, D., Taterka, H., and Tran, A. 2008. Influence of moisture on aggregation of protein during storage. Abstract book, Annual Meeting of USDA CREES PIs NRI Grants. New Orleans, LA.
• Labuza, T.P. 2008. 21 Material Science approach to the mechanism of food crispness. Marcel Loncin Award Symposium, 10th International of Engineering of Foods Congress, Valpariso, Chile
• Zhou, P., Liu, X., Tran, A., and Labuza, T.P. 2008. Effect of polyols on functionality and stability of whey protein based nutritional bars. Abstract book of IFT Annual Meeting, New Orleans, LA
• Liu, X., Labuza, T.J., Metzger, L.E. , and Labuza, T. P. 2008. Effect of processing aids on heat stability of protein-based beverages, Abstract book IFT Annual Meeting New Orleans
• Wang, N., Blasius, T., Diez-Gonzalez, F. and Labuza, T.P. 2008. Thermal and chemical inactivation of ricin in orange juice. Abstract book NCFD Annual Meeting Chaska, MN.

Progress 01/01/07 to 12/31/07

Outputs
OUTPUTS: This project is designed to study the effects of moisture content (or water activity, aw) and temperature on the growth or inactivation kinetics of pathogenic organisms, bioterror agents and degradation kinetics of health oriented functional ingredients (vitamins, nutraceuticals, protein) as a function of time in storage and undergoing Accelerated Shelf Life Testing (ASLT). The outcome is to help the food/drug and nutraceutical industry design both safe and effective products and understand the influence of the environment on the loss of efficacy of the product through using the aw stability map, shelf life plot and thermodynamic state diagram -glass transition line. Four trade journal articles were published, 2 by an editor (Food Technology 40K circulation) and 2 by us covering ASLT and aw principles. In the area of the inactivation kinetics of food papers were presented at the Inter. Assoc. for Food Protection meeting in Banff (> 1000 persons), at the NCFPD annual meeting (200) and at the IFT national meeting in Chicago (20,000 ) covering work on anthrax and ricin inactivation by heat and biocides and on Listeria growth kinetics based on time to detect for hot dogs, deli turkey and ham slices. Presentations were also made to MN meat processors and the MN House Bioscience and Innovation Research Comm. A public lecture on bioterrorism was given to > 200 persons at Gustavus Aldolphous College in MN. The chemical work has focused on understanding and mechanisms to prevent protein aggregation reactions in nutritional protein/energy bars. Such a reaction makes them hard and lose nutritive value. Presentations in this area were, a 2nd lecture as the Sigma Xi Distinguished Science Lecturer at Gustavus Adolphous (100 persons) , a presentation before the Industry Dairy Panel at Dairy Management Inc in Chicago (10 industry leaders), the invited Tanner lecture at the Chicago IFT Section (100 persons), the MN Particle Soc. meeting in St Paul, MN (40 persons), an invited speaker at the DMI/IFT Whey Symposium at the IFT annual meeting in Chicago (~300 persons) , as an invited plenary presentation at the Xth International Symposium on the Properties of Water (ISOPOW) attended by food and pharmaceutics scientists, physicists, physical chemists and plant biologists (200 participants) and a practicum for the Thai food industry on the influence of water on food stability, an acute problem in a humid country (>200 participants). In addition we have worked with several instrument manufacturers (Decagon - $50K instrument grant; Thermo Fisher $250K contract) to study moisture sorption and stability eg the latter on aflatoxin detection and deflection on whole grain and with an international medical device company on design of ASLT procedures. Another key output was the editing and writing a material science approach textbook on water activity in foods. Prior books in this area were symposium proceedings, with a text focused on microbial growth. PARTICIPANTS: Dr. Michael Uzzan, Post Doc Position, PhD Technion Israel - currently Tamara Foods Haifa Dr. Gerry Schamburger, Post Doc Position, PhD Food Science U of MN - currently Heinz Corp. Pittsburg PA. Dr Peng Zhou, Post Doc Position, PhD Cornell - currently Assoc. Prof Jiangnan Univ Wuxi China Dr Xiaoming Liu, Post Doc Position, PhD Washington State Univ. - currently Asst. Prof. Jiangnan Univ. Wuxi China Dr Sa Xu, Post Doc Position, PhD Iowa State - currently Land O'Lakes Co. Minneapolis, MN Dr Amit Pal, PhD student, U of MN. - currently Campbell Soup Campden, NJ Amy Tran, MS Student, U of MN, Food Science Heather Taterka, MS Student, U of MN Collaborators Dr Baram Ismail, Asst. Prof. U of MN, Food Science TARGET AUDIENCES: Nutraceutical Bar Manufacturers Dry inhalation Drug manufacturers Fluid Milk Processors Ready to Eat MeatProcessors USDA/FDA regulatory personnel

Impacts
This project is designed to study the effects of moisture content (or water activity, aw) and temperature on the growth or inactivation kinetics of pathogenic organisms, bioterror agents and degradation kinetics of health oriented functional ingredients (vitamins, nutraceuticals, protein) as a function of time in storage and undergoing Accelerated Shelf Life Testing (ASLT). The outcome is to help the food/drug and nutraceutical industry design both safe and effective products and understand the influence of the environment on the loss of efficacy of the product through using the aw stability map, shelf life plot and thermodynamic state diagram -glass transition line. In the area of the inactivation kinetics of bioterror agents we found that the normal 70 C, 15 sec HTST pasteurization regime is not adequate to inactivate anthrax spores or ricin to safe levels in milk contrary to some USDA published data. Use of biocides like hydrogen peroxide in combo with temperature treatments at 90C, capable of doing in most dairy plants, gives a reliable process to decontaminate the milk before environmental disposal, the process is not designed to make the milk edible. In 2006 the FDA and USDA proposed that shelf life dates placed on Ready to Eat foods like hot dogs could be based on growth of Listeria from below detection level to either the Time To Detect (TTD) or some higher level. Our study assessed the growth kinetics of 19 pathogenic strains and 4 were found to be the fastest. These were then inoculated on hotdogs, sliced ham and sliced turkey and held at 4 temperatures until TTD. A plot of log TTD as a function of temperature gave good parallel straight lines for each initial non-detectable low concentration. The work showed that, if temperature abused at > 7C, detectable and infectious levels of Listeria would occur before the labeled 90 day shelf life of the meats studied. The work on nutraceutical degradation in processing and storage showed that there is great differences in stability. The focus was a small 4 oz milk serving containing up to 5 different dietary supplements beneficial to human health. The findings were presented in the last year report and the paper was published this year. We are continuing to evaluate creatine stability in dry vs semi-moist vs liquids. Preliminary data suggests that the typical min max rate curve vs aw does not occur in liquid systems, this needs further study. Our main focus this year has been on protein aggregation reactions in the dry and semi-moist state leading to loss of nutritional quality and hardening of the protein bar making it too tough to chew. Our results with whey isolates parallel what has been found for dry inhalation protein drugs like insulin. During storage there is intermolecular disulfide bond formation which is the cause of the problem. Cysteine and glutathione which are reducing agents exacerbate the problem while hydrolyzed whey or glycerol reduces the hardening probably by lowering the glass transition temperature and thus becoming more plasticized. We are continuing to look for other means to slow the reaction.

Publications

• Xu, S., Labuza, T.P. and Diez-Gonzalez, F. 2007. Inactivation of Bacillus anthacis spores by a combination of biocides and heat in milk at pasteurization temperatures. National Center for Food Protection and Defense Annual Meeting.
• Zhou, P., Davis, L., Schamberger, G., and Labuza, T. 2007. Maillard reactions in high protein nutritional bars. Inter. Society for Maillard Reaction, Frankfurt, Germany.
• Labuza, T.P., Labuza, T.J., Labuza, K.M. and Labuza, P.S. 2007. Soft condensed matter: A perspective on the physics of food states & stability ISOPOW 10, Bangkok, Thailand.
• Labuza, T. and Zhou, P. 2007. Moisture-induced aggregation between protein molecules/particles in whey protein powders and protein bar model systems. Minnesota Particle Society, St Paul, MN.
• Labuza, T.P., Labuza, T.J., Labuza, K.M. and Labuza, P.S. 2007. Soft condensed matter: A perspective on the physics of food states and stability. IFT Tanner Lecture, Chicago, IL.
• Belcourt, L. and Labuza, T.P. 2007. Effect of raffinose on sucrose recrystallization and textural changes in soft cookies. J. Food Sci. 72(1):C65-C71.
• Diez-Gonzalez, F., Belina, D., Labuza, T.P., and Pal, A. 2007. Modeling the growth of Listeria monocytogenes based on the time-to-detect in culture media and frankfurters. Intl. J. Food Microbiology 113:277-283.
• Uzzan, M. and Labuza, T.P. 2007. Thermal and storage stability of nutraceuticals in a milk beverage dietary supplement. J Food Science 72(3):E109-117.
• Zhou, P. and Labuza, T.P. 2007. Effect of moisture content on the glass transition and protein aggregation of solid-state whey proteins and their hydrolysates. Jr. Food Biophysics 2: 2007 108-116.
• Xu, S., Labuza, T.P., and Diez-Gonzalez, F. 2007. Inactivation kinetics of avirulent bacillus anthracis spores in milk with a combination of heat and hydrogen peroxide. J. Food Protection 7(6):4479-4483.
• Labuza, T.P., Peng, Z., Davis, L. 2007. Whey to go: Application of whey proteins in nutrition bars J. of The World of Food Ingredients (Nov) pp.108-112
• Pal, A., Diez, F., and Labuza, T.P. 2007. Shelf life dating for seafood, time for safety-based approach. Global Aquaculture Advocate March/April pp. 32-35.
• Rahman, M. and Labuza, T.P. 2007. Water activity and food preservation, Chapter 20 In M. Rahman (ed.) Handbook of Food Preservation 2nd Edition. Taylor & Francis, Boca Raton, FL.
• Labuza, T.P. and Altunakar, B. 2007. Water Activity Prediction and Moisture Sorption Isotherms Chapter 5 In G. Barbosa-Canovas,A. Fontana, S. Schmidt, and T.P. Labuza (eds.), Water Activity in Foods: Fundamentals and Applications. IFT Blackwell Press. Ames, Iowa. pp 109-154.
• Labuza, T.P. and Altunakar, B. 2007. Diffusion and sorption kinetics of water, Chapter 9 In G. Barbosa-Canovas, A. Fontanna, S. Schmidt and T. Labuza (eds), Water Activity in Foods: Fundamentals and Applications. IFT Blackwell Press, Ames, IA, pp 215-238
• Barbosa-Canovas, G., Fontana, A., Schmidt, S. and Labuza, T.P. (co-editors) 2007. Water Activity in Foods: Fundamentals and Applications. IFT Blackwell Press, Ames, IA.
• Xu, S., Labuza, T.P., and Diez-Gonzalez, F. 2007. Inactivation of Bacillus anthracis spores by a combination of biocides and heat in milk at pasteurization temperatures. IAFP, Banff, Canada.
• Raabe, O.N., Labuza, T.P. and Diez-Gonzalez, F. 2007. Extraction of Bacillus anthracis spores from milk. IAFP, Banff, Canada.
• Labuza, T. and Zhou, P. 2007. Whey protein stability and functionality in bar applications (symposium). IFT Annual Meeting DMI/Dairy Foods Division, Chicago, IL.
• Zhou, P., Tran, A. and Labuza, T. 2007. Softening of whey protein based nutritional bars by whey protein hydrolysates. IFT Annual Meeting, Chicago, IL.
• Xu, S., Pal, A., Labuza, T.P. and Diez-Gonzalez, F. 2007. Thermal and chemical inactivation of ricin in milk. IFT Annual Meeting, Chicago, IL.
• Pal, A., Labuza, T.P. and Diez-Gonzalez, F. 2007. Predictive modeling of Listeria monocytogenes in ready-to-eat frankfurters for safety-based consume-by date labels. IFT Annual Meeting. Chicago, IL.
• Zhou, P. and Labuza, T.P., 2007. Moisture-induced whey protein aggregation in the protein/water model system. International Dairy Federation; Dairy Proteins. San Francisco, CA.
• Diez-Gonzalez, F., Labuza, T.P. and Raabe, O. 2007. Extraction of Bacillus anthracis spores and ricin from liquid foods. National Center for Food Protection and Defense Annual Meeting.
• Xu, S., Pal, A., Labuza, T.P. and Diez-Gonzalez, T.P. 2007. Thermal and chemical inactivation of ricin in milk. National Center for Food Protection and Defense Annual Meeting.

Progress 01/01/06 to 12/31/06

Outputs
The paper on nutraceutical stability during thermal processing and storage in a dairy beverage was accepted for publication in early 2007. DMI has published several press papers of the results and several companies are interested in creating such functional foods. This was given impetus after the 12/06 meeting on the proposed FDA regulations for functional foods. The paper on creatine degradation kinetics remains to be submitted. Work on anthrax inactivation kinetics in milk was completed and published including a paper on front face fluorescence (FFF) to measure heat damage during thermal processing and one on using green tea antioxidants to reduce such damage. The latter received wide spread publicity on several industrial website including Food Navigator. FFF is also being researched as a means for continuous 100% inspection of cereal grains for the presence of mycotoxins with the ability to divert contaminated kernels before they enter the food chain. Other continuing work is on rapid techniques to isolate spores from fluid foods and evaluation of the degradation kinetics of ricin in thermal processing of milk. The work on designing shelf life dates based on food safety using Listeria as the model pathogen continues. A general paper based on early results for modeling of time to detect as a model for food safety dating will appear in the Int. Jr. of Food Microbiology (07) and 4 presentations were made in Europe. We have identified 5 strains isolated from the major food outbreaks showing maximum growth rates and we are now testing them for growth rates on hotdogs, turkey deli meat and sliced ham. Following this we will test out prediction models and compare to sensory quality loss time. Work on the physics of hardening of marshmallows and the stickiness of hardball confections was presented at the IFT annual meeting and at Eurofoods Water meeting in Brussels. It was found that marshmallow hardening is solely a loss of moisture phenomenon, unlike the work done of soft cookies where it is the result of sugar crystallization causing loss of plasticizer and moisture redistribution. Impermeable foil laminate packaging will prevent this but at a cost. For hardball candy, temperature or humidity abuse to about 10 to 15 C above the Tg for the product will result in stickiness onset while at 50 degrees above Tg results in collapse and complete flow into one solid mass. At 65 C collapse and flow occurred at all humidities. Given that inside car temperatures, when left in the sun, can rise to 40 C. above the air temperature, such candies will not survive. Recent work on the mechanism of hardening of high whey content nutritional bars indicates that the mechanism consists on moisture redistribution in a few days after mixing followed by significant disulfide bond formation between protein chains and between chains of different particles causing network formation while the Maillard reaction is only significant if high fructose corn syrups are used. Means to reduce hardening are being evaluated.

Impacts
The study on degradation rates of dietary supplements during heat treatment and storage in a milk based drink designed for the elderly will be published in Jr. Food Science in March, 2007. This work has been cited in several trade journals and several companies interested in creating such products. Further work on thermal processing of milk has shown that addition of green tea antioxidants can reduce cooked flavor. Employing shelf life dating based on time to detection of a pathogen (in our case Listeria) was shown to be feasible and could be employed by the cured meat industry. Hardening of marshmallows was shown to be solely water loss which could be prevented by better packaging. Stickiness of hard candy and cough drops could be prevented by both humidity control (better packaging) and temperature control. However leaving such candies in a closed car in the sun will lead to unacceptable product. . The new work on prevention of hardening of protein bars to improve quality and nutrition will be expected to be implemented by industry this year.

Publications

• Leinen, K. and Labuza, T.P. 2006. Crystallization inhibition of amorphous lactose using raffinose. J. of Zhejiang University SCIENCE B Applied Physics and Engineering Sciences. 7(2):79-83.
• Xu, S., Labuza, T.P. and Diez-Gonzalez, F. 2006. Thermal inactivation of bacillus anthracis spores in milk. J. Applied Environmental Microbiology. 72(6)4479-4483.
• Sherwin, C. and Labuza, T.P. 2006. Beyond water activity and glass transition: a broad perspective on the manner by which moisture can influence reaction rates in foods. In Water Properties of Food, Pharmaceutical and Biological Materials. M. Pilar, et.al. ed. Taylor and Francis NY. (ISBN 0-8493-2993-0).
• Schamberger, G. and Labuza T.P. 2006. Use of Front-face fluorescence for assessing thermal processing of milk. J. Food Sci. 72(2):C69-74.

Progress 01/01/05 to 12/31/05

Outputs
The degradation of dietary supplements functional for the elderly in a milk based drink for the elderly was evaluated. This included chondroitin sulfate, glucosamine, lactoferrin, creatine, lutein and soy isoflavones. The products were pasteurized at 72 C to 138C in a Microthermics unit followed by storage for up to 7 months at 4 to 45 C. The nutraceuticals were measured using new HPLC methods adapted to milk except for Lactoferrin which was measured by immuno-diffusion. The study demonstrated the technical feasibility of using a milk beverage as a convenient and palatable vehicle for delivery of nutraceutical compounds. Stability was mostly limited by either degradation of the bioactive component or interactions with milk components such as browning or co-precipitation. Isoflavones and chondroitin sulfate were stable in all tested thermal treatments and during storage for 7 months at 21 C (RT) and 4 months at 45 C. Glucosamine enriched milk can be pasteurized, but it destabilized the milk proteins if heated at greater than 80 C. Storage stability of glucosamine followed apparent first and zero order kinetics in early storage with rate constants that fit the Arrhenius equation (Log shelf life vs temperature) up to 45 C where degradation leveled off. Assuming 20% loss allowed, end of shelf life would be about one month at RT or nearly two months if refrigerated. Lactoferrin was reasonably stable in pasteurization but denatured at UHT conditions. No degradation of lactoferrin was found during storage at room temperature or lower during 5 months. Creatine was satisfactorily stable in thermal treatments but degraded rapidly in storage. These nutraceutical compounds can be used to design milk beverage dietary supplements but a concentration of up to 20% excess may be required for some of them. The study of the mechanism of hardening of whey protein based bars which leads to unacceptability continues. We have found that substitution of 20% soy isolate significantly decreases the hardening. Using chemical methods, it seems that the reaction is disulfide bonding between proteins causing aggregation and thus the bars hardens. We are now looking at other ways to stop this reaction. A new technique, front face fluorescence (3F), which directly measures the development of off flavor precursors without sample preparation was tested. Fluorescence was both linearly related to hydroxy methyl furfural formation as well as OD at 450 nm. We are continuing to evaluate this with a sensory panel, testing milk pasteurized at 120 C for 10 sec. Recently the National Advisory Committee on Microbiological Criteria for Foods proposed that shelf life of ready to eat refrigerated foods be based on pathogen growth. Thus we began to study the growth rate characteristics (time to detect, lag time, and log rate constant of 19 strains of Listeria monocytogenes at 5 temperatures in the refrigerated and abuse range. We developed a unique method to evaluate the time to initial detection at 1 CFU/25 g. These data will form an algorithm for setting a safety based expiration date as a function of time-temperature in distribution, ie. the goal of NACMCF, USDA and FDA.

Impacts
The study demonstrated the technical feasibility of using a milk beverage as a convenient and palatable vehicle for delivery of nutraceutical compounds.

Publications

• Labuza, T.P., Diez-Gonzalez, F., and Belina, D. 2005. Alternatives to Safety Based Shelf Life Dating. Food Protection Trends 25(1) 53-56
• Hyman, C. and Labuza, T.P. 2005. The brittle-ductile transition of an amorphous food system. J. Drying Tech. 23 (4): 871-886
• Hyman, C. and Labuza, T.P., 2005. Correlating perceived crispness intensity to physical changes in an amorphous snack food. J. Drying Tech. 23(4):887-906
• Roe, K. and Labuza, T.P. 2005. Glass transition of Amorphous Trehalose-Sucrose systems. J. Food Properties 8(3):1-16
• Kempf, T., Levine, L. and Labuza T.P. 2005. Wheat flour dough flow visualization during a sheeting process. Cereal Foods World 50(6): 293-372.
• Heldman, D.R., Clausi, A.S., Kokini, J., Labuza, T.P., Rizvi, S.S. and Swartzel, K.L. 2004. IFT Research Needs Report, Food Technology. 58(12):pp.32-36.
• Davis, Laurie. 2005. Evaluation of the hardening of whey protein based energy bars. M.S. Thesis, University of Minnesota. 420 pp.
• Belcourt, Laura. 2005. Effect of raffinose on sucrose recrystallization and textural changes of soft cookies. M.S. Thesis, University of Minnesota. 305 pp.

Progress 01/01/04 to 12/31/04

Outputs
The degradation kinetics of nutraceuticals in a milk base drink for the elderly is being done. Focus is on stability of dietary supplements important to the elderly including chondroitin sulfate, glucosamine, lactoferrin, creatine, lutein and soy isoflavones during pasteurization and UHT processing in a Microthermics unit and storage for 16 weeks + at 25 to 45C. Concentration was measured using new HPLC methods adapted to milk. Lactoferrin which was measured by immuno-diffusion. Soy isoflavones and chondroitin sulfate were stable during processing and storage (16 weeks). Glucosamine and lactoferrin caused precipitation of milk protein when processed at >100C but not at 72C. For storage evaluation, these were cold-sterilized (Millipore membrane),and to the UHT milk. Glucosamine/milk was unstable with a shelf life of 2 months at 4 C and 1 month at 21C, while lactoferrin was stable at 4 and 21C but degraded rapidly at 45C. Creatin was stable to pasteurization with a loss of <5% occurred at UHT conditions. Degradation of creatine was also evaluated at different water activities and pH values of 4, 7, and 10. A combination of all the nutraceuticals was evaluated for the possibility of interactions that may affect stability. A formulation of such a beverage was prepared by aseptically mixing a sterile solution of glucosamine and lactoferrin with UHT heated pre-mix of chondroitin sulfate, isoflavones, and creatine in skim milk. The resulting product was stored in 4, 23, and 35 C. Shelf life is under evaluation including a lutein study. Since lutein is known to be light sensitive, this is also being evaluated. One concern is the off flavor developed as the result of non-enzymatic browning at UHT processing temperatures followed by refrigerated storage. Means to suppress this using natural plant antioxidants (eg epicathecin) and other potential inhibitors (NaHSO4, EDTA, cysteine, etc. is ongoing. A new fluorescence technique which measures the fluid directly without prep is being evaluated. This has potential for keeping milk acceptable if a more robust process is needed to destroy potential bioterror agents. We are developing models for the food distribution chain needed to track product flow under a grant from the Dept. of Homeland Security. In other studies we finished the modeling of the temperature profile in a two-fluid heat exchanger, needed for kinetics of degradation. We also determined the time to detection of listeria on Ready to Eat meats (hot dogs) as a function of temperature which allows an algorithm to be established for setting an expiration date as a function of time-temperature in distribution. The use of raffinose to prevent the hardening of soft cookies in storage is also being evaluated in comparison to trehalose which slows hardening. Raffinose shows significant reduction of crystallization of sucrose in a cotton candy matrix at 5% (7 days vs 3 days at 33%RH but may be less protective in a cookie matrix.

Impacts
The FDA has acknowledged in their proposed GMPs for dietary supplements that processors must guarantee label values for their ingredients. Our work focuses on the use of a easy to drink (swallow) milk based beverage made stable at room temperature by ultra high thermal processing (UHT). We have set the stage for the types of test procedures needed for evaluation of both processing and storage conditions on stability for key nutraceuticals. Early work has shown that in thermal treatment (pasteurization) and storage soy isoflavones, creatine and lactoferrin are very stable. We have also developed a mathematical model to predict the temperature profile under UHT, which was previously not solved for. Thus we can now evaluate the overall degradation kinetics at each stage of the heating treatment. As a collaborating project our studies of UHT can be used to predict inactivation of bioterror agents and we have, with Penn. State found some means to eliminate the non-enzymatic browning off flavors produced under UHT processing conditions. Several food grade antioxidants have the ability to block the reaction. Finally our work on increasing the shelf life of soft cookies has now indicated that although sugar crystallization is a key step in hardening, it is more the redistribution of moisture throughout the cookie that is the key to keeping it soft. Trehalose seems to function as a viscosity control agent limiting hardening much like high fructose corn syrup while raffinose is a direct crystallization inhibitor

Publications

• Labuza, P.S. and Labuza, T.P. 2004. Cotton candy shelf life. J. Food Processing & Presv. 28:274-287.
• Uzzan, M. and Labuza T.P. 2004. Critical issues in the R&D of soy isoflavone enriched foods and dietary supplements. J. Food Science 69(3): CRH 77-86.
• Uzzan, M. and Labuza, T.P. 2004. Temperature profile within a counter current flow plate and frame heat exchanger. J. Food Science 69 (9): E433-440.
• Labuza, T.P., Belina, D., and Diez-Conteras, F. 2004. Food safety management in the cold chain via expiration dating. In Cold chain management. B. Pederson and B. Kunz (Editors) Univ. Bonn, Germany. http://www.iaph.uni-bonn.de/coldchain/
• Labuza, T.P, Roe, K., Payne, C. Panda, F., Labuza, T.J., Labuza, P.S. and Krusch, L. 2004. Storage stability of dry food systems: influence of state changes during drying and storage. In Drying 2004. M. Silva and S. Rocha (editors). pp. 48-68 Ourograf Grafica Campinas, Brazil (ISBN # 85-904573-1-1) http://www.feq.unicamp.br/~ids2004/
• Labuza, T.P. and Myers, T. 2004. Perishables and RFID Part 1. Food Production Management 127(6): 6-8, 24
• Labuza, T.P. and Myers, T. 2004. Perishables and RFID Part 2. Food Production Management 127(9): 8-10, 24
• Belina, Dan. 2004. Time To Detection of Listeria below the Detection Limit. M.S. Thesis, U. of Minnesota, 176 pp.
• Sholl, Jeff. 2004. Modeling of the Food Distribution system for analysis of a bioterror event. Ph.D. Dissertation, U. of Minnesota, 225 pp.

Progress 01/01/03 to 12/31/03

Outputs
Under the prior aims of the project the influence of temperature and moisture content on lactose crystallization was studied During storage, whole milk powder (WMP) can adsorb moisture and undergo time-temperature dependent changes such as caking, sticking, and collapse followed by the crystallization of lactose, which are undesirable leading to browning and insolubility. When WMP was stored at room temperature at water activities in the range 0.11-0.5, crystallization was seen only seen above aw 0.33 at room temperature which is just at the glass transition temperature, Tg, for that moisture. The effect of temperature upon the time to crystallization (tc) below 0.33 was determined during isothermal storage (25 to 155 C). At aw 0.22 (6% moisture), lactose crystallized in one hour at 50 C, while at the lower moistures it occurred in one hour at 71 (aw 0.05) and 120 C (aw 0.11). The rate of crystallization increased as T-Tg increased, but the rate of crystallization was significantly different at the same T-Tg for moisture content. An Arrhenius model (ln k vs 1/T) fit the data at each moisture content better than a linearized WLF (T-Tg) model. The latter model as the driving factor for crystallization is flawed as moisture plays a critical role. Powder X-ray diffraction (XRD) was used to determine the rate of crystallization of lactose in WMP at abuse humidities (64%, 73%, and 77%RH). The crystallization of lactose began after 24 hours and ended after 96 hours for the WMP stored at 64%RH, while at 73% and 77% RH, it began immediately and ended sooner. Only alpha mono-lactose crystallized at the two higher water activities. A zero-order model for all three water activities appropriately described the crystallization of lactose in WMP. In a continuation of the soft cookie project it was found that trehalose at 5% as a substitute for high fructose corn syrup reduced crystallization by increasing the local viscosity as did the HFCS. Under the new research direction, after securing a microthermics unit, it was found that soy isoflavones added to milk were not destroyed over a wide range of UHT and conventional pasteurization conditions. A storage study is currently in progress. In addition we were able to derive mathematical equations that describe the time-temperature profile in both co- and counter-current heat exchangers including plate and frame type, something never solved to date and needed for kinetics experiments to compare predicted to actual losses in processing. In other studies we developed a method to determine the time to detection of listeria on Ready to Eat meats (hot dogs) as a function of temperature which allows an algorithm to be established for setting an expiration date as a function of time-temperature in distribution. Mathematical models for flow of dough through calendaring rolls, were developed allowing one to establish reduction ratios and speeds to prevent tearing in industrial operations . A method to determine pressure inside a High Pressure Processing (HPP) unit was found and we showed that with hams, the profile is not isostatic, thus more time is required in processing to get a desired pathogen kill.

Impacts
The FDA has acknowledged in their proposed GMPs for dietary supplements that processors must guarantee label values for their ingredients. We have set the stage for the types of test procedures needed for evaluation of both processing and storage conditions. Early work has shown that in thermal treatment (pasteurization) soy isoflvones are stable.

Publications

• Refereed Papers: Sherwin, C and Labuza, T.P. 2003. Role of moisture in Maillard Browning Reaction Rate in intermediate moisture foods: Comparing solvent phase and matrix properties. J. Food Sci. 68(2):558-94.
• Minerich, P. and Labuza, T.P. 2003. Development of a pressure indicator for high hydrostatic pressure processing. J. Innovative Food Science and Emerging Technologies 4(3): 235-243.
• Anderson, B.E., Shimoni, E. and Labuza, T.P. 2003. Degassing kinetics of carbon dioxide in packaging of fresh roasted ground coffee. J. Food Engineering 59(1):71-78.
• Manzacco, L., Nicoli, M.C., and Labuza, T.P. 2003. Study of bread staling by X-ray diffraction analysis. Italian Food Technology: Processing and Packaging XXXI(3):17-23.
• Book Chapters: Taoukis P. and Labuza T.P. 2003. Time-temperature Integrators, In Novel Food Packaging Techniques. ed., R. Ahvenainen, Woodhead Publ. Denver, CO. pp 104-126.
• Krusch, L. 2003. The kinetics of lactose crystallization in whole milk powder. M.S. Thesis, University of Minnesota. 225 pp.
• Puhek, T.J. 2003. Texture changes in soft cookies and the effect of Trehalose. M.S. Thesis, University of Minnesota. 308 pp.
• Kempf, T.P. 2003 Dough sheet model verification by flow visualization. Ph.D. dissertation, University of Minnesota. 349 pp.

Progress 01/01/02 to 12/31/02

Outputs
Research continues on loss of quality and shelf life of foods as related to processing, storage and packaging with a focus on sugar crystallization phenomena. For example soft cookies (like chocolate chip style with 25% sucrose or more) become firm in texture (crumbly to hard) in 1 to 6 days after manufacture, causing loss of acceptability and increasing food waste. However there are several commercial methods used to increase shelf such as by adding glycerol or high fructose corn syrup (HFCS) but these have lower sensory appeal and don't increase it much beyond 3 to 4 months while 6 to 9 months would be desirable. The basic mechanisms for loss of textural acceptability is not agreed upon. Sucrose is in a rubbery amorphous (non-crystalline state like cotton candy) in the cookie right after baking. This amorphous phase with water plasticizes the structure making it soft. Crystallization during storage is one of several hypotheses as the cause of firming during room temperature storage, yet no one has measured and confirmed this directly. The idea is that the rate of firming is due to sugar crystalline bridges forming throughout the cookie perhaps linked by the starch granules. The hypothesis for use of HFCS then is that it inhibits crystallization of sucrose, thus reducing the rate of firming. Using a Siemens 5005 powder Ray diffractometer, scanning electron microscopy and a texture instrument, we were able to show a high correlation between % sucrose crystallized and firmness. However at different initial sugar contents the amount of sugar needed to crystallize decreases as the initial sugar content is lowered, opposite of one expects based on bridge formation. In addition microscopy showed no bridges were formed. This suggests that sucrose crystallization is only a first step in firming, the main factor being that as it crystallizes, the water held by the sugar initially is lost to the starch granules reducing the amount of plasticizer volume needed to keep the cookie soft. We showed that the softening mechanism for HFCS was not as a crystallization inhibitor but that it increased the plasticizer volume which keeps the cookie soft. Studies with another purported crystallization inhibitor, trehalose, showed the same effect. Further work is continuing with other inhibitors such as raffinose to see if we can improve shelf life. With cotton candy we showed that moisture gain from the air was the key factor that causes collapse and graininess, a result of sucrose crystallization. Collapse and graininess are the major problems for commercial distribution. At 32%RH, cotton candy collapses in 3 days at room temperature, in one day at 45% RH and 2 hours at 75% RH. We have developed a mathematical model that predicts end of shelf life due to crystallization as a function of relative humidity. This model can be used commercially to determine the required plastic packaging material needed for a given shelf life. In addition that 30% w/w trehalose/sucrose cotton candy crystallized slower most likely due to a viscosity increase.

Impacts
We investigated fundamental physical properties as related to the crystallization of sucrose which cause textural problems leading to consumer food unacceptability. Knowledge of the reasons for sugar crystallinity and it's control will lead to a significant improvement in shelf life of soft cookies, a significant need in the bakery field. These same principles applied to cotton candy can help create a system to distribute the product without collapse and thus create a new successful business.

Publications

• Leman, M. L. 2002. Evaluation of the Rotovisco to measure melting characteristics of cheese. Plan B Masters Project, University of Minnesota, St. Paul, pp. 50
• Manzocco, L., Nicoli, M.C. and Labuza, T.P. 2002. Impiego dell analisi di diffrazione dei raggi X (XRD) per lo studio dei fenomeni di raffermamento del pane. Techica Molatoria 53(12):1208-17.
• Manzacco,L., Nicoli, M.C. and Labuza, T.P. 2002. Study of bread staling by x-ray diffraction analysis. Italian J. of Food Science 14(3):235-246.
• Minerich, P.L. 2002. The design and testing of a high hydrostatic pressure indicator for processing of meat. PhD Thesis, University of Minnesota, St. Paul. pp. 490
• Panda, F.A. 2002. The physical chemistry of texture changes in soft cookies. MS Thesis, University of Minnesota, St. Paul. pp. 365
• Sherwin, C.P. 2002. A molecular mobility approach to describing the role of moisture in the Maillard reaction rate. PhD Thesis, University of Minnesota, St. Paul. pp. 231
• Sherwin, C., T. P. Labuza, A. McCormack and B. Chen. 2002. Cross-polarization/magic angle spinning NMR to study glucose mobility in a model intermediate-moisture food system. J Agr Food Chem 50:7677-7683
• Vanchy, P. 2002. Improving the stability of spray dried whole milk powder. MS Thesis, University of Minnesota, St. Paul. pp. 186
• Labuza, T.P. and Schmidl, M.K. 2002. Medical foods in issues in food safety. R. Schmidt Editor J. Wiley and Sons. NY
• Shimoni, E., Dirks,E. and Labuza, T.P. 2002. The relationship between final popped volume of pop-corn and thermal physical parameters. Lebensmittlen Wiss u Tech 5:93-98

Progress 01/01/01 to 12/31/01

Outputs
Research continued to focus on shelf life predictions as related to storage and packaging. With respect to bakery type foods, the physical chemistry of quality loss in soft moist cookies was continued. These cookies quickly (12 to 48 hours) become hard after baking, causing loss of acceptability and increasing food waste. After baking the sucrose is in the amorphous rubbery state as indicated by its position above the glass transition temperature vs. moisture curve (Tg Vs m) as measured by differential scanning calorimetry. Sucrose crystallization begins immediately after baking and leads to the toughening of the cookie, following the same kinetic rate as pure amorphous sucrose published in 1956. This indicates that the cookie ingredients offer little ability to reduce diffusion in the cookie. Importantly, the individual sucrose crystals themselves do not make the cookie hard since they are not interconnected. A Siemens 5005 powder Xray diffractometer was used to determine % sucrose crystallinity over time which closely correlated to increased firmness as measured by a texture instrument. It is clear that the loss of moisture from the amorphous sucrose is picked up by the gluten and the starch granules. This moisture loss reduces the plasticizing effect of the sugar-water matrix and leads to firming. Thus crystallization inhibitors should increase shelf life. However the common use of high fructose corn syrup only reduces the initial rate over a six day period. At 21 days both show the same degree of crystallinity and are unacceptable texturally. Use of sugar laser scanning NMR techniques indicates in model systems that local diffusion is very limited however for crytallization rotational freedom is likely enough for it to occur. Thus current work is focussing on use of trehalose which has been touted to be an effective inhibitor. If enough inhibitor is added crystallinity should be prevented and a six month shelf life is possible. The other possibility is the use of a non-aqueous plasticizer such as glycerol to keep the cookie soft but it might also increase the rate of crystallization. These same principles of crystallinity kinetics above the Tg curve were applied to storage stability of cotton candy. Cotton candy if unprotected quickly (in hours) collapses if exposed to temperatures above 30 C or humidities above 45%. It was shown that this product, which is glassy and brittle when made, needs a very high moisture barrier moisture film to maintain it's glassy state which gives it acceptable texture. A gain of 2 grams per 100 grams in storage leads to collapse and crystallization which occurs in three days at 33%RH. At 3% moisture the product also collapses above 40 C. The stability is directly predicted by the Tg curve with respect to moisture gain and temperature increase. Current work will also look at the substitution of trehalose for sucrose.

Impacts
We investigated fundamental physical properties as related to the crystallization of sucrose to investigate textural problems which lead to consumer food unacceptability. Knowledge of the reasons for sugar crystallinity and it's control will lead to a significant improvement in shelf life of soft cookies, a significant need in the bakery field. These same principles applied to cotton candy can help create a system to distribute the product without collapse and thus create a new successful business

Publications

• Schmidl, M.K. and Labuza, T.P. 2001 Medical Foods in Current Issues in Food Safety R. Schmidt and Gary Rodrick Editor J. Wiley and Sons. NY
• Labuza, T.P. 2001 Nutraceuticals: GMPs, Safety, Stability and Date Coding in "Examining the Science Behind Nutraceuticals". G. Gold Editor AAPS Press, Va.
• Cardelli, C and LabuzaT.P. 2001 Weibull Hazard Analysis to the Shelf Life of Fresh Roasted Ground Coffee. Lebensmittlen Wiss u Tech 34(5): 273-278
• Duyvestyen, W., E. Shimoni and T.P. Labuza. 2001. Shelf life of fluid milk using the Weibull Hazard Method. Lebensmittlen Wiss u Tech 34:143-148
• Laaksonen, T, Y. Roos and Labuza T.P. 2001 Effects of sucrose, NaCl, water content and arabinogalactan on relaxation in wheat dough as measured by DMTA J. Food Properties 4: 311-325
• Laaksonen, T, Y. Roos and Labuza T.P. 2001 Comparisons of the use of desiccators with or without vacuum for water sorption and glass transition. Int. J Food Properties 4: (In press)
• Shimoni, E. , E. Anderson. and Labuza, T.P. 2001 Reliability of Time Temperature Indicators under Temperature abuse . J. Food Science 66(5): 1337-1340.
• Shimoni, E., Dirks, E. and Labuza, T.P. 2001 Evaluation of the physics of pop-corn popping. Lebensmittlen Wiiss und Tech (In press)
• Book Chapters Schmidl, M . K. and Labuza, T.P. 2001 US legislation and functional food claims in Functional foods G. Gibson and C. Williams Editors Woodhead Publ. Co. London
• Sablami, S., Rahman, S., and Labuza T.P. 2001 Chapter A.2.3 Measurement of water activity using the isopiestic method in " Current Protocols in Food Analytical Chemistry. R. Wrolstad ed. J. Wiley & Sons New York
• Rahman, S., Salami, S N. Guzanni, Labuza, T.P. and P.P.Lewicki 2001 Chapter A2.4. Direct manometric measurement of water activity In "Current Protocols in Food Analytical Chemistry" R. Wrolstad ed. J. Wiley & Sons New York
• Labuza, T.P., Cardelli, C., Andersen, B., and Shimoni, E . 2001 Physical chemistry of carbon dioxide equilibrium and diffusion in tempering and effect on shelf life of fresh roasted ground coffee. Proceedings 19th ASIC R. Liardon Editor ASIC Press. Paris France
• Labuza, T.P. 2001 Nutraceuticals: GMPs, Safety, Stability and Date Coding 2001 Proceedings of pre-Internet Conference 11 World Congress of Food Science and Technology KoSFoST Soeul Korea pg 410
• Szybist , L. , Labuza, T.P and J Peck. 2001 Perishable refrigerated products and home practices survey. Working paper # 2001-4 The Sloan Foundation Food Retail Industry Research Center Univ. of Minnesota
• Books: TP Labuza and Lynn Szybist 2001 Open dating of Food. Food and Nutrition Press 239 pp Trumbull, CN.

Progress 01/01/00 to 12/31/00

Outputs
Research focused on loss of quality and shelf life of foods as related to processing, storage and packaging. We studied the physical chemistry of staling of French style bread and soft moist cookies. Both products quickly become firm, staling, after manufacture (12 - 48 hr, causing loss of acceptability and increasing food waste. The basic mechanisms for loss of textural acceptability is not agreed upon but there are several methods to produce soft products but of lower sensory appeal. In cookies, the high sugar level prevents starch gelatinization and there is little gluten development. Thus, sucrose crystallization in storage is one of several hypotheses as the cause of staling during room temperature storage, yet no one has measured and confirmed this directly. Using a Siemens 5005 powder Xray diffractometer, we were able to (1) create a quantitative Xray intensity curve vs % sucrose crystallinity previously only done for pure amorphous-crystalline drug systems, and (2) show that there is a high correlation between % sucrose crystallinity and firmness. Both firmness and crystallinity followed the expected first order curve and the sucrose crystallization matched the kinetics of the Avrami equation. With French-style bread, staling is hypothesized to be due to starch crystallization because of the water content and low sugar content. We: (1) produced a standard curve for starch Xray crystal intensity as a function of crystallinity which has not been done before and (2) showed that the XRD patterns correlated very well with textural firmness. In retail bread, the loss of water through the packaging material also contributes to firming due to a transformation on the state diagram from a rubbery to a glassy state. These results indicate the direction to reduce loss of textural acceptability should focus on prevention of crystallization. In other areas we (1) evaluated the reliability and predictability of a new commercial Time-temperature Integrator (3M Fresh Check) and found it to be highly reliable for use on refrigerated food (time to loss of acceptability); (2) showed that a potentially improved method for predicting shelf life based on lipid oxidation has severe limitations; (3) determined the spore death kinetics in a peroxide-vacuum sterilizer; (4) completed a study on diffusion kinetics of carbon dioxide loss from fresh roasted ground coffee; and (5) evaluated the water binding properties of two nutritionally functional ingredients (arabinogalactan and inulin). In addition several book chapters, and a book on the product development and regulation of functional foods world wide were completed along with a revision of a book on methods of water sorption determination.

Impacts
Some bakery goods become unacceptable after processing because of physical changes that occur during storage. This resaerch has focused on the mechanism that leads to loss of crispness and on the hardening of bread and soft cookies. The results will enable processors to increase shelf life therby reducing significant food waste.

Publications

• Mauer, L.J., Smith, D.E., Labuza, T.P. 2000. Effect of water content, temperature and storage on the glass transition, moisture sorption characteristics and stickiness of beta-casein. Intl. J. Food Properties 3: 233-248
• Mauer, L.J., Smith, D.E., Labuza, T.P. 2000. Water vapor permeability, mechanical and structural properties of edible of beta-Casein films. J. Food Engineering. In press.
• Schaller-Polvony, L.S., Smith, D.E., Labuza, T.P. 2000. Effect of molecular weight and water content on the moisture sorption characteristics of commercial inulin sources. Int. J Food Properties 3:173-192.
• Shimoni, E. and Labuza, T.P. 2000. Degassing kinetics and sorption equilibria of carbon dioxide in fresh roasted ground coffee. J. Food Engineering 23:419-436.
• Cardelli, C and Labuza, T.P. 2000. Application of Weibull Hazard Analysis to the shelf life of roasted and ground coffee. Lebensmittlen Wiss u Tech 33: In press.
• Duyvestyen, W., Shimoni, E. and Labuza, T.P. 2000. Determination of the end of shelf life of fluid milk using the modified Weibull Hazard Method. Lebensmittlen Wiss u Tech 33: In press.
• Shimoni, E. and Labuza, T.P. 2000. Analysis of predictive microbiology methods for the prediction of growth of pathogens below the detectable level. Trends in Food Science and Technology 11(X) In press.
• Labuza, T.P. 2000. The search for shelf life, Food Testing Analysis, April/May 6(2):26-35.
• Labuza, T.P. 2000. Functional foods and dietary supplements: Safety. good manufacturing practice (GMPs) and shelf life testing. In essentials of functional foods. M.K. Schmidl and T.P. Labuza, Editors, Aspen Press, London
• Schmidl, M.K. and Labuza, T.P. 2000. Nutrition and health claims in food labeling. J. Ralph Blanchfield, Editor, Woodhead Publ. Co., London
• Schmidl, M.K. and Labuza, T.P. 2000. U.S. legislation and functional food claims in functional foods. G. Gibson and C. Williams, Editors, Woodhead Publ. Co., London
• Bell, L.N., and Labuza, T.P. 2000. Practical aspects of moisture sorption isotherm measurement and use. 2nd Edition, AACC Eagan Press, Eagan, MN.
• Schmidl, M. and Labuza, T.P. 2000. Essentials of functional foods. Aspen Press, London.
• Hyman, C. 2000. Correllating preceived sensory crispness intensity changes to physical changes of an amorphous food system. Ph.D. Thesis, University of Minnesota, St. Paul. pp. 399.
• Schaffer, S.M. 2000. Kinetics of vacuum H2O2 inactivation of bacterial spores in a vacuum proxide. Ph.D. Thesis, University of Minnesota, St. Paul. pp. 133.
• Fitch, N. 2000. Evelauation of a new method to predict oxygen uptake as a function of oxygen partial pressure. M.S. Thesis, University of Minnesota, St. Paul. pp. 172.
• Ju, G. 2000. Functional food regulations and product technology in Japan vs. the U.S. M.S. Thesis, University of Minnesota, St. Paul. pp. 140.
• Anderson, B. 2000. Kinetics of carbon dioxide diffusion loss from fresh roasted ground coffee. M.S. Thesis, University of Minnesota, St. Paul. pp. 176.

Progress 01/01/99 to 12/31/99

Outputs
Temperature is a major factor in determining the shelf life of refrigerated and frozen foods.In 1996 the federal government (FDA, USDA) concluded that temperature abuse is a major concern in distribution. FDA published a Federal Register notice (11/22/96)to evaluate various alternatives to ensure better temperature management. FSIS has commented that the suspect hot dogs in the Bil Mar Foods Listeria deaths were close to end of shelf life and were mostly likely temperature abused. Most manufacturers of refrigerated products use an open date (sell by, use by, best if used buy) in order to facilitate distribution but if abused, the date is misleading and leads to a false sense of security. Currently open dating is left up to the states, except for infant formula. The US Congress's Office of Technology Asessment (OTA) examined the potential for mandatory Federal legislation in 1979 but never acted on it. We compiled all State and Territory laws on food dating on the web (www.trfic.umn.edu) along with call for a mandatory Federal dating law. We cataloged all products with dates in a large supermarket and found varied and confusing usage unfriendly to the consumer. OTA in 1989 concluded an on package tag indicating accumulated temperature abuse be designed to facilitate better distribution practices by showing a color change before the date stamped if temperature control was not sufficient. Three such products are commercially available. Through efforts of The Retail Food Industry Center (TRFIC), Cub Foods introduced hamburger with such tags in 1999. In a survey of consumer knowledge/use of open dating, we found that pre-cut salads and pasteurized milk were the categories with the greatest spoilage (19 and 7% respectively) before the stamped date. More than 50% of consumers had home refrigerator temperatures > 41 F, the maximum recommended for potentially hazardous foods. Food safety scares also induced 50% of them to reject the indicted food or all brands in the category. We found that sensory crispness of dry snack foods is related to basic physical properties (glass transition , Tg, and brittle-ductile transition) as a function of moisture and temperature. Crisp materials become soggy at a specific combination of moisture-temperature values. We are evaluating enzymatic protein aggregation means to raise Tg so as to create a crisper cookie. The influence of moisture on the Tg of popping corn was studied. A 3 X greater volume occurs if pre-humidified to 62% RH which is related to the thermal breaking properties of the pericap. The factors controlling the desorption rate and equilibrium of carbon dioxide from fresh roasted ground (FRG) coffee were studied. After roasting, coffee has 10 mL of CO2/gram, thus expensive films that have a pressure relief valve are required for flexible packages unless the gas is first desorbed. Sorption equilibrium follows Henry's Law but there is a significant amount of non-equilibrium gas entrapped in the coffee structure. Diffusivities for CO2 useful in the design of desorption equipment have also been measured.

Impacts
Federal mandatory open dating of refrigerated foods will reduce consumer confusion on shelf life. Use of a time-temperature integrating tag may induce better temperature management and reduce food borne illness. Refrigerator appliance manufacturers need to make more intelligent devices to indicate temperature as it is a problem in the home. Sensory crispness of dry snacks is controlled by glass transition thus processing changes would lead to a crisper product.

Publications

• DeSobry, S.A. , Netto, F.M. and Labuza, T.P. 1999 Preservation of B-carotene from carrots . Critical Reviews in Food Science and Nutrition. 381-396 pp.
• DeSobry, S.A., Netto, F.M. and Labuza, T.P. 1999 Influence of maltodextrin systems at an equivalent 25 DE on B-carotene loss during spray drying and storage. J. Food Proc. Preserv. 23:39-55
• Rahman, M. and Labuza, T.P. 1999 Water Activity and Food Preservation Chapter 11 in "Handbook of Food Preservation". pg 339-382 . Marcel Dekker New York
• Caldwell, E.F, Johnson, L.E. and Labuza, T.P. 1999 Fortification and Preservation of Cereals in Breakfast and How They are Made. 2nd Edition. E.F. Caldwell ed. Eagan Press MN

Progress 01/01/98 to 12/31/98

Outputs
The application of glass transition phenomena is being explored to understand the crisp to soggy transition in dry snack products when they gain moisture. At a given plasticizer content (moisture) there exists a temperature (Tg) above which diffusion and mobility of molecules is significantly enhanced. Several methodologies including dynamic mechanical thermal analysis (DMTA) , measurement of molecular mobility by nuclear magnetic resonance (NMR), electron spin resonance (ESR), and movement of radiolabled molecules and degree of sugar recrystallization by X-Ray diffraction are being used. In addition a sensory technique to evaluate crispness is being developed to determine if loss of crispness always occurs at some isoelasticity line at a constant T above Tg. This would allow one to manipulate ingredients to raise or lower Tg and thus control texture of finished cereal products. Glass transition can also apply to hardening of soft cookies during storage, fracture of pasta, sugar glazing of cereals and volume expansion of pop corn. A new method is being developed to shorten the time needed to measure oxidative rancidity of cereals (oxygen uptake over time) from 1 1/2 years to several months. This would greatly enable optimization in the selection of packaging. Fresh roasted ground (FRG) coffee contains 8% by weight of carbon dioxide out of the roaster, requiring tempering for 24 to 48 hrs before sealing in flexible packages otherwise they burst. The loss rate as a function of temperature and humidity is being studied to shorten tempering time and optimize use of flexible films. The Mac software packages developed in 1993 for sorption properties and kinetics of shelf life have been transformed into Excel spreadsheets. There is great demand for these programs from labs all over the world to enable analysis of shelf life parameters of foods, seeds and biologics.

Impacts
(N/A)

Publications

• DESOBRY, S.A. , NETTO, F.M. and LABUZA, T.P. 1998. Comparison of spray drying, drum drying and freeze drying for encapsulation and stability of Beta-carotene. J Food Science 24: 1158-62
• NETTO, F.M, DESOBRY, S. and LABUZA, T.P. 1998. Effect of water content on the glass transition, caking and stickiness of protein hydrolysates. J Food Properties 1: 141-161
• HYMAN, C. and LABUZA, T.P. 1998. Moisture migration in multidomain systems. Trends in Food Science and Technology 9(2): 47-55
• DAVIES, C.G.A., NETTO, F., GLASSENAP, N., GALLAHER, C.M., LABUZA, T.P. and GALLAHER, D.D. 1998 The Development of Maillard reaction products during storage of protein isolates. J Agric Food Chem. 46(7):2485-2489
• DAVIES, C. , KAANANE, A. LABUZA, T.P., A. MOSKOWITZ, A.J. and GUILLAUME, F. 1998. Evaluation of the acyclic state and effect of solvent type on mutarotation kinetics and Maillard browning rate of glucose and fructose. In 'The Maillard Reaction in Foods and Medicine' Editors: O'Brien, J., Nursten, H., Crabbe, M.J. and Ames, J. Royal Soc Of Chemistry, London. P. 166-171
• TAOUKIS, P. and LABUZA, T.P. 1998. Chemical time-temperature integrators as quality monitors in the chill chain. Proceedings of the International Symposium Quimper Froid 1997. Predictive Microbiology of chilled foods. June 16-18, (1998). IIR and European Commission, COST 914.
• TUTUNCU. M.A. 1998. The Effect of Geometry on the Effective Diffusion Coefficient in Ready-To-Eat Cereals. Ph.D. Thesis. Univ. of Minnesota, St. Paul. P. 411.

Progress 01/01/97 to 12/31/97

Outputs
The study of the shelf life of fresh roasted ground coffee was completed. We developed the first mathematical model which successfully integrates the effect of variable oxygen partial pressure (pO2), moisture content and temperature (T) during distribution of a dehydrated food in a flexible package film that is permeable to both oxygen and moisture. This model predicts the influence of lipid oxidation rate (k) on sensory shelf life as determined by the Weibull Hazard method. Temperature dependent data was modeled by the classical Arrhenius approach (ln k vs 1/T) and water activity (aw) dependence by the Ln k vs aw approach. Oxygen reaction rates were measured by GC. Over 76 diffferent conditions of pO2, aw and T were utilized. The influence of temperature on the growth of microbes and the sensory shelf life of specialty milk products as determined by the Weilbull Hazard method has also been completed. It was found that sensory shelf life was independent of the microbial level. Several time-temperature integrating devices were tested which allow for prediction of end of shelf life for variable T distribution. The stability of beta-carotene was evaluated after encapsulation by three drying methods, with drum drying being the best. The glass transition (Tg) state diagram and caking stability was determined for casein, whey and fish protein hydrolysates. These gave a lower Tg than for monomeric sugars indicating a potential for use in product development.

Impacts
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Publications

• NETTO, F.M., DESOBRY, S. and LABUZA, T.P. 1997. Effect of water content on the glass transition, caking and stickiness of protein hydrolysates. J. Food Properties 1:(in press).
• DESOBRY, S., NETTO, F.M. and LABUZA, T.P. 1997. Influence of maltodextrin systems at an equivalent 25DE on beta-carotene loss during spray drying and storage. Jr. Food Procs. and Preserv. 25:(in press).
• HYMAN, C. and LABUZA, T.P. 1997. Moisture migration in multidomain systems. Trends in Food Science and Technology 9:(in press).
• DESOBRY, S.A., NETTO, F.M. and LABUZA, T.P. 1997. Preservation of beta-carotene from carrots. Critical Reviews in Food Science and Nutrition (in press).
• CARDELLI, C. Ph.D. 1997. Shelf Life of roasted ground coffee as a function of water activity, temperature and oxygen. Ph.D. Thesis. Univ. of Minnesota, St. Paul. 343 pp.
• TUTUNCU, A. and LABUZA, T.P. 1997. Effect of Geometry on Moisture Transfer Diffusion Coefficient. J. Food Eng. 28:1-15.
• Z. LONG, CHEN, P.L., RUAN, R.R. and LABUZA, T.P. 1997. Nuclear Magnetic Resonance Studies of Water mobility in bread during storage. Lebensmittlen Wiss. und Tech. 30:178-183.

Progress 01/01/96 to 12/30/96

Outputs
The flow properties of a cereal based product before and after baking, influences consumer textural acceptance. Perceived texture is determined by the temperature/moisture position on the state diagram (glass transition temperature plot of Tg vs moisture content), ie rubbery (soft, elastic) or glassy (crisp, brittle). Texture is related to the visco-elastic properties of the constituent polymers and sugar monomers, including amylose, amylopectin, gliaden and glutenin in flour as well as any added sugars. Dynamic mechanical thermal analysis (DMTA) and differential scanning calorimetry (DSC) studies are continuing on various cracker formulations. A fermented cracker has a completely different Tg compared to a chemically leavened one, due to complete metabolism of low MW sugars and peptides, thus maintaining cracker crispness up to 20%(wb) moisture. We are evaluating NMR as a technique for Tg measurement. For maltodextrins it shows similar results as compared to DSC. Further work will be on composite foods. We have completed a study of the glass transition properties of several dairy and fish hydrolyzed proteins and evaluated the changes in solubility during storage. We are trying to determine whether there is a pH drop in model systems as the aw is decreased using pH indicators. We completed an evaluation of a new aw measurement component for Dunmore/Pope sensor type instrument that helps to speed measurement (5 - 10 min vs 18 hours).

Impacts
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Publications

• NIKOLADIS, A. and LABUZA T.P. 1996. Glass transition curve of crackers and relationship to gluten. J Food Science 61(4):803-806 NIKOLADIS, A. and LABUZA, T.P. 1996. (DMTA) as an appropriate method for measuring the glass transition temperature of a composite food product. J. Thermal Analysis 47:1314-1328 TUTUNCU, A. and LABUZA, T.
• P., 1995. Effect of Geometry on Moisture Transfer Diffusion Coefficient. J Food Eng . 28:1-15 NIKOLADIS, A. 1996 Glass transition phenomena and the texture of cereal products. M.S. Thesis. Univ. of Minnesota, St. Paul. 180 p.

Progress 01/01/95 to 12/30/95

Outputs
The flow property of a cereal based dough and the finished texture after baking,are an important process control parameter (e.g. size) and consumer acceptance characteristic. This is defined by the position on the state diagram (glass transition temperature or Tg vs moisture content) for the dough during processing and as the finished product, i.e. rubbery (soft, elastic) or glassy crisp, brittle). The flow and texture are controlled by the visco-elastic properties of the constituent polymers as defined by the Williams Landau Ferry free volume theory. These include amylose, amylopectin, gliaden and glutenin in flour. Using Dynamic Mechanical Thermal Analysis (DMTA) and Differential Scanning Calorimetry (DSC) it was found that the glass rubber transition in a low sugar biscuit formula is controlled by the combined gluten fraction, however the curve is depressed due to the interactive effects of sugar as described by the Gordon-Taylor equation. The starch component was not controlling as verified by the fact that the dough and finished cracker showed the same Tg curve and the starch was not gelatinized during baking. A high sugar formula and the influence of added dairy based proteins on the Tg curve is now under investigation. In addition a modulated temperature profiling DSC has been purchased and is being investigated as to its applicability to define the Tg curve. =Tg curves of other foods including chewing gums and fresh roasted ground coffee have been made.

Impacts
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Publications

Progress 01/01/94 to 12/30/94

Outputs
A new instrument, a Dynamic Mechanical Thermal Analyzer with 1000 x greater sensitivity than DSC for measuring thermo-elastic events was tested to see if it can be used to measure glass transitions in finished food products. The main use of the DMTA has been to measure such properties for synthetic polymers and some food polymers such as starch and gluten. This instrument uses a dynamic oscillator to measure elastic moduli. For polymers, at constant plasticizer content, a dramatic drop in elastic moduli occurs at the glass temperature (Tg) and causes materials to become soft. Thus it was proposed that loss of crispness of cereal snack foods is a result of the transition from an amorphous glassy state to a rubbery state as it gains moisture and becomes soft due to a decrease in elastic moduli. A prior theory of this lab concluded that crispness was lost due to easier sliding of polymer chains above a certain moisture content. The instrument was found to be very capable of measuring the Tg curve of a finished food product, namely a baked cracker. In addition the glass transition curve for the unbaked dough was superimposable over the curve for the finished product. The combined curve comes closest to that of the curve for gluten compared to the other cracker components. These results suggest that gluten is the main polymer that controls texture and crispness and that modification of the gluten could be used to improve texture and reduce packaging requirements.

Impacts
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Publications

Progress 01/01/93 to 12/30/93

Outputs
Caking, powder stickiness and collapse are physical properties related to the position (glass transition temperature (T(subscript g)) vs. moisture) on the state diagram which determines stability. Differential scanning calorimetry is a relatively new method to obtain this. The state diagram was determined for three high fat commercial infant formula products and compared to conventional sticky point and caking methodologies which have large variabilities. Tg was found to be an easily measured predictor of physical collapse phenomenon as well as the critical % ERH at room temperature at which caking and sticking occurs. The results indicate that maltodextrins can be used to reduce caking of high sugar-high fat dairy powders. The effect of degree of glassy vs rubbery state on oxidation of the whey protein encapsulated butter fat was evaluated during storage at several relative humidities (%RH). Dairy protein encapsulated high butter fat powders were produced by spray drying at several temperature conditions. For protein-fat mixtures like infant formula, when stored at different %RH, the glass transition state had little effect on oxidative stability. The oxidation pattern followed the typical pattern of a rate increase above and below the monolayer moisture suggesting that the free volume dependence of oxygen diffusion is not limited in the glassy state.

Impacts
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Publications

Progress 01/01/92 to 12/30/92

Outputs
Further work continued on the measurement of pH of actual reduced-moisture foods. It was clearly shown that the use of a wide surface membrane sensor as in the CARDY pH meter gave the most accurate estimate of the pH in the dry state. This will be invaluable in the prediction of stability of dry and semi-moist foods. Work on the influence of glassy vs rubbery state of encapsulation polymers used to create spray dried encapsulated lipids indicates that stability can be achieved in the rubbery state if the physical collapse that occurs forms a surrounding envelope with low oxygen permeability. This might be useful in storage of spray dried flavors. Work on edible moisture barriers indicates that at a level of 25-30% fatty acid, minimum water vapor transmission rate (WVTR) is achieved and that stearic acid gives the lowest permeability of the commercial fatty acids. Casein based films could be formed with good physical strength but exhibited poor WVTR. These films might have application for non-food use where biodegradability is a need such as in diaper closure tape, package inner liners and decorative ribbon. Browning degradation of dry and semi-moist foods by the Maillard reaction was found to be controlled by the physical state (rubber vs glass) of the system, and both WLF and Arrhenius kinetics were applicable at all moisture contents. Thus at least three temperatures are needed in making shelf life predictions at these moisture contents.

Impacts
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Publications

Progress 01/01/91 to 12/30/91

Outputs
Various methods (dilution of 1 g solids with 9 mL water, dilution with 99 mL water, wetting surface surface with a few drops of water, Gran plot method-serial dilution with extrapolation to zero added water, microtip probe, and Cardy surface meter) used to evaluate the pH of dry and semi-moist food systems at reduced water activity (a(w)) were compared on several model systems made to different a(w)s. Results were compared to the pH predicted using degradation rates of acid/based catalyzed chemical markers (aspartame, ampicillin) in the systems corrected for the effects of humectants on the pH of different buffers at reduced water content. The study showed that the pH of systems in which water has been removed decreases due to the increase in the hydrogen ion activity but the change is different than predicted by the Debye Huckle equation corrected for change in dielectric constant and ion concentration. The Cardy Surface pH meter gave pH values closest to that found from the degradation rates of chemical markers at low a(w). There is no theoretical direct prediction of the drop in pH with a decreasein a(w). The hydrogen ion activity increased by 0.5 to 1.5 times (increase in concentrations of 3 to 30 times).

Impacts
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Publications

Progress 01/01/90 to 12/30/90

Outputs
MacIntosh computer programs with complete graphics for calculation of the water activity (a(subscript w )) of a complex food based on system composition, for isotherm calculation and plotting with BET and GAB parameters, and for reaction rate calculations have been developed. A dimensionless number (L#) to mathematically test the efficacy of a moisture barrier to extend shelf life was derived based on film permeance and the diffusivity of the food. It was found to apply well and thus will save development time for edible barriers. A method to evaluate diffusivity of water in dense substances was developed so as to study edible barrier efficacy. The study of the loss of aspartname in dry and intermediate moisture model systems is being used to evaluate what the actual pH of limited aqueous systems is, since different degradation products form at different pH values. Wet gels systems buffered to pH 5, when brought to lower moisture contents, give reaction rates as if the pH was at 3. The log rate vs pH function does not follow theoretical predictions of a ten fold increase per pH unit. Buffers themselves go to a lower pH as the system is reduced in moisture, indicating that the nature of the condensed phase is complex and related to changes in the ionic strength and dielectric constant. Adding salts and humectants to buffers give conflicting effects on pH reduction at reduced water content with some increases and some decreases.

Impacts
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Publications

Progress 01/01/89 to 12/30/89

Outputs
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Impacts
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Publications

Progress 01/01/88 to 12/30/88

Outputs
The Decagon CX-1, new water activity device, was evaluated. It operates on the principle of dew point measurement in the head space above a 1-8 g sample and can measure a(w) under 6 minutes. The device gave good reproducibility in the 0.6 to 0.98 range. It was found that there was no difference in a(w) between the surface and the core of surface mold ripened cheeses. A new dimensionless number was developed and successfully tested which calculates the relative resistance of the package film to that of the internal resistance of the food to moisture transport. This allows evaluation of the use of simple analytical models to predict gain or loss or moisture during storage. A mathematical analytical model was successfully derived using the GAB isotherm equation for prediction of moisture gain/loss. This allows calculations over the full isotherm rather than just the linear part used in previous analytical solutions. The GAB model was also successfully incorporated into a solution for moisture transfer between multicomponent foods of varying a(w) and with the external environment. The properties of chocolate as a moisture barrier were determined. A 6 mil layer of chocolate has the same resistance to diffusion of water as a 1 mil polyethylene film. A storage study of the drying out of individually packaged sliced cheese indicated that the oiling out of the cheese reduced the film moisture permeance significantly, thereby increasing shelf life.

Impacts
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Publications

Progress 01/01/87 to 12/30/87

Outputs
The moisture barrier properties of chocolate coatings were evaluated. Moisture permeability (k/x) was assessed over a range of relative humidity (32-85%). Moisture transport is controlled by the amount and degree of fat crystallinity. As %RH increases k/x increases as with cellulosic films, however, even at 80% RH, the k/x of a typical chocolate coating is equivalent to that of a 1 mil thick polyethylene film. The moisture permeability is 5 times less depending on the way the chocolate film is set on the surface. Chocolate also shows significant hysteresis in its moisture sorption as a function of water activity (a(W)). A new model gel system was developed to test the usefulness of chocolate under unsteady state conditions. A numerical solution to the packaging mass transfer equation was found for the GAB isotherm, which previously required numerical integration. Results agreed very well with prediction. Initial studies show that: cysteine is effective as an inhibitor of non-enzymatic browning as a function of temperature and a(w); other enzymes can be used to inhibit the rate of polyphenol-oxidase at reduced a(w); there is a rapid loss of aspartame at low a(w); Ionic strength and a(w) significantly affects the rates of cyclization of reducing sugars; a new model to predict the freezing point depression of ice cream mixes as a function of the a(w)-moisture curve for each component has been developed; A a(w) stress can possibly cause elicitation of secondary metabolism end products in plant tissues bio-cultures.

Impacts
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Publications

Progress 01/01/86 to 12/30/86

Outputs
Soluble and insoluble fiber fractions of 43 fruit, vegetable, cereal and gum sources were measured by TDF and NDF methods. Soluble fiber content of F and V sources did not correlate with water binding capacity (WBC). WBC of F & V sources were similar (3 g water/g fiber) using a dialysis method. Cereal-fiber binds less water (1-2 g water/g) while gums showed 5-12 g water/g fiber. Thus WBC is not a good index of the physiological response of fiber in relation to gut transit time and stool volume since cereal-fiber shows the best results. Other WBC methods showed that higher a(w) gives higher WBC, explained by a thermodynamic model. A new method can evaluate the relative importance of the major resistances to moisture transport for packaged dry foods. A dimensionless number, L#, that relates external (film) and internal (food) resistance was introduced. L# provides a quantitative criterion for the analytical method which predicts moisture gain or loss. This was verified for packaged corn flakes, dates and wild rice while predictions for cheese slices were poor because oil from the cheese changed film permeability. Moisture permeability of chocolate was evaluated for potential as an edible barrier. It is equivalent to polyethylene in moisture resistance. The finite element model accurately predicted moisture exchange between two systems. A literature review showed concern for the use of humectants to lower a(w).

Impacts
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Publications

Progress 01/01/85 to 12/30/85

Outputs
Water binding property studies of food fibers were expanded to include 42 fruit,vegetable, gum and cereal fiber sources. Water binding was measured by liquid water suction at a water activity of 1.0 and by a second method designed to simulate the osmotic conditions of the gut (280 mOsm). As expected, WBC under physiological conditions was lower by 2 to 10 times. The method used involved dialysis against polyethylene glycol and had high reproducibility. Using these conditions, the WBC of some vegetable fibers exceeds that of cereal fiber by 4 to 10 times (e.g., squash 12 g/g solids versus wheat bran 1 g/g solids). This is mainly attributable to the pectin content of the dietary fiber. Several methods of dietary fiber analysis were applied to the fiber sources, including the newly accepted AOAC method. With soluble fiber present, the values for any one fiber could vary by +/- 40%, possibly due to presence of starch and protein. These results could not explain the differences in WBC among fiber sources. Further work is planned. A finite element program was developed to predict the rate of exchange of moisture between mixed foods of different water activities. The method required development of a sorption technique to measure effective vapor diffusion coefficients. Over 15 foods were measured and used in the model, which gave excellent prediction of equilibrium time in mixed foods such as white/wild rice, cereal/raisin, and trail mixes of nuts, fruits and cereals.

Impacts
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Publications

Progress 01/01/84 to 12/30/84

Outputs
The water binding properties (WBC) of 15 vegetable and fruit fiber sources were evaluated on the Bauman device and by a new dialysis technique simulated by the human gut. Acorn squash fiber and oinion fiber have the highest WBC while potato has the poorest (a 12 to 5 fold respective difference). Most fruit and vegetable fiber sources have a better WBC than cereal fiber which is opposite to gut transport rates in the literature. Clinical work is needed to verify this. Three water activity (a(w)) devices for measuring water binding were evaluated. Results showed that the instruments (impedance and dew point) require calibration curves and that the calibration varies with the measurement temperature. Manufacturer suggested equilibrium times of < one hour are too short, although practical results could be obtained in two hours. Water binding by measurement of sorption isotherms was done in conjunction with the European Economic Community COST 91 project recommendations. A practical workbook for doing isotherms was published. A three week humidification time was adequate to achieve moisture gain/loss to within less than a 5% error (in most cases < 1%) of a six month humidification. Mathematical analysis of the moisture/a(w) curves showed that the GAB three parameter isotherm (Guggenheim-Anderson-deBoer) was the best fit for over 90% of the foods tested or published in the literature.

Impacts
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Publications

Progress 01/01/83 to 12/30/83

Outputs
Previous work showed that the Baumann Capillary Suction-Water Uptake Device (BSD) gave the best prediction of the ability of a food gum soruce to bind and hold water in a practical application, namely to maintain juiciness in a low fat cooked meat emulsion. Besides gums, food fiber sources such as bran are used extensively for water binding as well as for potential health benefits, because of their ability to increase intestinal motility. The water binding of 15 food fiber sources was measured by the BSD method, by determination of the sorption isotherm, by freezing point depression, and by the standard AACC swelling-centrifugation technique. The fiber sources bind about 10X less water than gums, on the average. No correlation could be obtained between the BSD and AACC techniques, probably due to the difference in the mechanism of water uptake in each device. Fruit fibers (apple, citrus pulp) bound 2X as much water as cereal bran sources. Rice bran bound 30% more than wheat bran, 100% more than oat bran and 300% that of corn bran. These results were used to make recommendatons on fiber choice for a human clinical study on gut motility. A two-year study evaluating new water activity (A(W)) measurement devices was completed. For both precision and accuracy in product development as well as to meet regulatory guidelines, the Beckman-Sina and the Kaymont-Rotronics impedance sensing devices were found to excellent.

Impacts
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Publications

Progress 01/01/82 to 12/30/82

Outputs
Water uptake data on the Baumann capillary device has been collected for a series of nine common food gums including plant exudates, modified starches and seaweed extracts. In addition, similar data has been collected for 12 types of dietary fiber sources such as soy and wheat brans. Information on water binding was also measured by osmometry and sorption isotherms. No correlation could be found between the latter two techniques and the Baumann uptake value. Four of the gums including the best (xanthan), two intermediate (carrageenan), locust bean) and the worst (low methoxy pectin) as measured by the Baumann were used to make a low fat hot dog. These were then tested for water binding by expressible moisture. Excellent correlation in rank was found between the hot dog water binding and the Baumann values. This suggests that application of Raoult's law and water activity lowering cannot predict gum functionality in a meat system. A similar test will be made with a cheese system. The fiber data will be used to choose four fibers to test in a human feeding trial to measure stool bulk and mineral absorption. Present data shows that the food fibers are much poorer water binders in general than the gums using the Baumann results. Of all the fibers apple pulp fiber can be considered to be the best based on the Baumann results. In the case of food fiber, a good correlation of water binding with the biophysical lowering of water activity was found.

Impacts
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Publications

Progress 01/01/81 to 12/30/81

Outputs
The water binding properties of eight common food gums, starches and fibers wereevaluated by measuring the sorption isotherm, water activity lowering by cryoscopic osmometry, water binding by the AACC soak/centrifugation method and water uptake rate and maximum retention using a capillary suction device. Several other devices, which can be used to measure water binding by water activity lowering, have been obtained and were evaluated in terms of their accuracy and precision. One of these, an electric absorption resistance device, has proven to be very accurate and will be utilized as well to measure the water binding of the finished food products when the functionality of food macromolecules is evaluated. The present results suggest that of the macromolecules teted, low methoxy pectin (a fruit gum) and carrageenan (a seaweed gum) have the best water binding and water holding capacity, but no theoretical relation has proved valuable in connecting the two properties. The AACC procedure was found to be totally unacceptable for the gums. Of interest is the fact that the water uptake rate does not correlate with the maximum water uptake. Thus, in processing a low-fat product, those macromolecules which take up water rapidly may be more desirable if process times are short such as in meat emulsion making, whereas those with slow uptake but hold more water may be desirable in low-fat cheese manufacture where the process time could be longer.

Impacts
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Publications

Progress 01/01/80 to 12/30/80

Outputs
The effect of water activity (a(w)) on the quality of several foods was investigated in the dry and intermediate moisture range. Whey powders were unexpectedly found to undergo the maximum extent of browning and protein quality loss at low a(w) (0.44). This maximum in rate was caused by the physical shift of amorphous lactose to the crystalline state resulting in trapped water pools as evidenced by Scanning Electron Microscopy (SEM). Reaction rates were faster in sealed vs. open systems since in the latter the water diffuses out of the pools more rapidly. The a(w) corresponding to the loss in crispness of dry snacks was determined to be about 0.3 to 0.45. This is similar to the range in which starches can undergo crystalline changes but SEM could not show this. A cryoscopic osmometer was used to evaluate the water binding properties of solute/carrageenan systems which can form gels. Sugar causes an interaction effect which lowers the a(w) more than carrageenan or sugar alone, which is due to cooperative hydrogen bonds. Overall the cryoscopic results showed that although gels can hold over 50-100 times their own weight, the reduction in water activity is minimal and thus their use as humectants for IMF products is negated. Riboflavin was found to be stable during storage of pasta in the dark, whereas about 25-50% loss of thiamine occurs above an a(w) of 0.4 and 30 degrees C.

Impacts
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Publications

Progress 01/01/79 to 12/30/79

Outputs
Analysis of enthalpy/entropy compensation phenomena did not provide a useful theoretical tool to predict the effect of water activity on rates of chemical reactions. The interaction of water binding agents with solutes was investigated in the high water activity range by use of a cryoscopic osmometer, which shows results comparable to the suction pressure method. As ionic and non-ionic solutes are added to gels, interactive effects occur which depress water activity greater than expected. Results indicate a cooperative effect of ionic and hydrophilic water binding which increases with concentration thereby facilitating stability of intermediate moisture foods. The rate of browning of whey powders in the IMF range shows a maximum of much lower water activity (0.44) than expected. Collapse as indicated by scanning electron microscopy results in local regions of available water thereby facilitating browning. Rate of quality loss is about the same for hygroscopic vs. non-hygroscopic whey although the latter contains only 5% amorphous lactose initially. Shelf life plots indicate that whey powder should be stored below 30 C and 40% RH to insure against rapid lysine loss if held in water permeable packages.

Impacts
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Publications

Progress 01/01/78 to 12/30/78

Outputs
In a continuation of the work of water binding by macromolecular gelling agents it has now been found that the previously proposed three dimensional capillary network is not necessary. Analysis of the Flory Huggins interaction parameter Chi, of the spin-lattice relaxtation time T(1) by pulsed nuclear magnetic resonance and by X ray diffraction, shows that for the four gels investigated (K-carrageenan, agarose, gelatin and potato starch) the binding of water in dilute gels can be predicted solely on the basis of 1) hydrogen bonds, 2) electrostatic bonding and 3) hydrophilic forces. At a certain break point concentration as measured by the water suction pressure curve the gels undergo a transformation from a dilute random solution with few junction points to an oriented capillary matrix with specific junction points. Above that breakpoint increased water binding occurs due to entrapment of water within the structure, with carrageenan being the strongest water binder and gelatin and starch the poorest. A strong association was made between macromolecular structure, numbers of available hydrophilic groups and ionic group content and the binding of water. The results were further confirmed in a dye diffusion study in the gels.

Impacts
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Publications

Progress 01/01/77 to 12/30/77

Outputs
A study of the mechanism by which macromolecular food ingredients affect water binding through gelation has continued. Water is held in these gels primarily by a capillary suction mechanism, the strength of which can be predicted by a new equilibrium technique developed in our lab. X-ray diffraction and electron microscopy confirm the capillary theory. NMR and dye diffusion results show that this water does not have a reduced water activity(a(w)). These results illustrate the complexity of the water binding process. Further refinement of the manometric procedure for a(w) measurement has been completed and tested against other devices. It proves to be the best method available for a measurement. The study of microbiological activity in intermediate moisture food (IMF) systems shows that Staphylococcus aureus nuclease production (an indicator of toxin) falls off sooner than does growth, indicating higher a(w) limits can be used. This reduction is a function of a(w) and type of humectant. Ethanol, in low concentration (1 to 4%) inhibits S. aureus above its minimum growth a(w) and thus is a useful relatively non-toxic humectant. A combination of glycerol/propylene glycol proved to be very efffective in preventing undesirable fermentation of colostrum milk which can be used in calf feeding programs. Application of reduced a(w) techniques was used to produce a shelf stable IMF processed cheese food with acceptable quality after 6 months at room temperature.

Impacts
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Publications

Progress 01/01/76 to 12/30/76

Outputs
The effect of humectants on the rate of non-enzymatic browning in intermediate moisture food (IMF) systems was studied. Liquid glycols decrease the browning rate at high water activity (a(w)) but shift the maximum to lower a(w). Maximum browning rate occurs at 3:1 molar ratio of reducing sugar to protein (lysine); however rate of reaction does not follow total available lysine content of protein. Analysis of a(w) measurement devices shows the VPM method to be most accurate (plus or minus 0.003 a (w) units) and the Abbeon device to be useful for quality control (plus or minus 0.01 units). A six month shelf life study of IMF processed cheese shows excellent stability microbiologically and organoleptically at a(w) 0.94 to 0.91. Preliminary studies show ethanol at 2 to 5% by weight to be an excellent microbial inhibitor at reduced a(w) in both liquid and solid food systems. Method of addition and sequence of addition of multiple liquid and solid humectants to IMF systems does not affect a (w) lowering ability. The Ross equation based on independence of each humectant in the total water phase accurately predicts final product a(w). A new method was developed to measure water binding of gel forming macromolecules based on suction potential. A break in suction potential at 10% gelatin is confirmed by NMR, viscoelastic, and diffusion measurements.

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Progress 01/01/75 to 12/30/75

Outputs
REACTION STABILITY - The loss of quality of casein by non-enzymatic browning in an intermediate moisture food (IMF) system was studied using DNFB - available lysine and a biological system Tetrahymena pyriformis W, (protozoan). In early stages of browning, lysine becomes chemically unavailable but initial results show the lysine to be available to the organism suggesting that the aldose-aminereaction is reversible. Studies of IMF systems undergoing oxidation were made to test the effectiveness of antioxidants as a function of temperature. It was found that at IMF water activity BHA is the best antioxidant whereas in dry systems the BHA/BHT combination is more effective. Other antioxidants were onlyslightly effective. Extrapolation of oxidation rates from 45 C down to 25 C cannot be made because of a change in free radical mechanism. Sorption water holding capacity of twelve widely used humectants was determined. In addition, equations were developed and tested for predicting a lowering ability of these humectants in food. A successful method was found using the Ross equation or theslope/ratio method. This was used to develop an IMF process change.

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Publications

Progress 01/01/74 to 12/30/74

Outputs
Stability of intermediate moisture foods (IMP) was studied with respect to non-enzymatic browning (NEB), loss of vitamin C, lipid oxidation, and microorganism viability. Increasing reducing sugar to lysine content ratio increases rate of NEB up to a 3:1 molar ratio. Above that browning rate is constant. Proteins react at different browning rates due to structural differences. Casein and fish protein concentrate react at similar rates; soy, whey, egg and gluten proteins react slower in decreasing order. Lysine loss follows the same pattern. Addition of liquid glycol humectants lowers browning rate and lysine losses significantly. Lowering processing temperatures maximizes storage browning rate. Vitamin C destruction increases with moisture content to 20% water content and then remains constant. Storage of IMP in a palladium catalyst scavenger pouch with a N(2)/H(2) atmosphere decreases vitaminC loss and completely prevents rancidity. Accelerated test procedures were developed for predicting IMP shelf-life. Microbial growth below water activity (a(w)) 0.84 is limited to molds. Challenge procedures were developed. Growth limits vary with food composition and pH. Significant antimycotic activity at pH 5.6 was found for 2% propylene glycol, 2% butane diol, 2% glycerol, methyl/propyl paraben, and 0.002% pimaricin.

Impacts
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Publications

Progress 01/01/73 to 12/30/73

Outputs
An isopiettic method was developed to determine water activity in the high A(w) range where other techniques are not accurate. This was tested on several high moisture foods and found to be very effective. Lipid oxidation studies show that water at high enough concentration will dilute the aqueous phase decreasingthe effective metal catalyst concentration and thereby reducing oxidation rate. A longer shelf life for intermediate moisture foods (IMF) can thus be obtained if the moisture content is in this range. At lower A(w), moisture-increases increased oxidation due to reduction in mobile phase viscosity and thus increased mobility. NMR studies of the model systems (Horticulture--Frost Hardness Lab) verified the differences in phase viscosity and bound water with respect to water sorption hysteresis. Vitamin C destruction showed no change inkinetic mechanism with respect to A(w). The rate of loss was directly proportional to moisture content as based on phase dilution. Thus the sorption hysteresis effect rates at the higher moisture and for desorption systems because of lower viscosity. Studies of antimycotics have shown that chemicals other than K-sorbate can be used effectively in IMF even at high pH. Studies have also verified the change in minimum growth A(w) with hysteresis in other food systems.

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Publications

Progress 01/01/72 to 12/30/72

Outputs
Mechanism of action of water at high water activity (A(w)) was studied relative to lipid oxidation. Oxidation rate increases with moisture. Proved conclusively that concentration of trace metals present in aqueous phase as wellas swelling of the network which exposes new catalytic sites both contribute to catalytic activity. For low metal systems, in IMF fruit and vegetable products,increasing A(w) increases development of rancidity. However, at a certain A(w),rancidity rate decreases due to catalyst dilution by increased water content. For meat products where metal concentration is high and bound, usually to protein, A(w) increases swelling making more metals available; thus oxidation rate increases. Antioxidants such as BHA and BHT are not very effective inhibitors at high A(w) since they operate only in lipid phase. However, water soluble chelating agents such as EDTA and citrate are effective since they decrease activity of trace metals. Addition of nitrate directly or indirectly from vapor space accelerates oxidation due to change in redox potential. Preliminary studies on vitamin stability (B(1) and C) show high destruction rates at high A(w) with rate directly dependent on moisture content. Studies ofmicrobial growth confirmed that minimum A(w) concept must be modified with respect to sorption hysteresis.

Impacts
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Publications

Progress 01/01/71 to 12/30/71

Outputs
A new sorption manometric measuring device was designed and built in order to beable to measure water activity accurately in the intermediate moisture range. This was tested using standard salt solutions and found to be highly reliable ascompared to the old hygrometer method. A pork IMF system was designed for five relative humidities and prepared by both a desorption and adsorption technique. Glycerol level was kept constant in all systems. The systems were inoculated with four species of microorganisms and stored at 25 C. In all cases the limiting A(w) was higher for the adsorption systems indicating that growth is not just A(w) dependent. It is postulated that the effect is not dependent on glycerol level as previously felt. A dog food system was studied at A(w)=0.8 for mold inhibitor effectiveness. Glycerol itself has no effect whereas potassium sorbate gives the best protection at the 0.1% level. 1,3-butane diol has some mold inhibitor effect at the 3% level. A system has been designed to study the independent effects of A(w) and water content on lipid oxidation. A non-swelling cellulose base material was used to give the product some texture. A 10% difference in moisture content can be achieved in the IMF range so that the effect of viscosity on the system can be evaluated. The oxidation rates arebeing followed by Warburgs and by peroxide determination.

Impacts
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Publications